Title of Invention

A NUCLEIC ACID OR FUNCTIONAL FRAGMENT THEREOF ENCODING PROTEINACEOUS MOLECULE AND A METHOD FOR PRODUCING A FLAVOUR EMPLOYING THE SAME

Abstract The present invention relates to the filed of genetic engineering of flavor, fragrance and biocontrol agent development. More specifically it relates to a process for production of natural flavors, fragrances or bio-control agents by the control of one or more genes implicated in that process. The invention provides an isolated or recombinant nucleic acid or functional fragment thereof encoding a proteinaceous molecule essentially capable of flavor, fragrance and/or bio- control agent synthesis when provided with a suitable substrate under appropriate reaction conditions. The invention further provides a nucleic acid or functional fragment thereof encoding a proteinaceous molecule essentially capable of synthesizing at least a monoterpene alcohollinalool when contacted with geranyl diphosphate (GPP) and/or at least a sesquiterpene alcohol nerolidol when contacted with farnesyl diphosphate (FPP) under appropriate reaction conditions.
Full Text

Title: Isoprenoid Synthases
The invention relates to the field of genetic engineering of flavor, fragrance or bio-control agent development. More specifically it relates to a process for production of bioactive isoprenoid compounds by the control or modulation of one or more genes implicated in that process.
Isoprenoids are the largest and most diverse group of plant secondary compounds. At least 20,000 isoprenoids have been described and without doubt many more will be discovered in the future. By definition isoprenoids are made up of socalled isoprene (C5) units. This can be recognized in the number of C-atoms present in the isoprenoids which usually can be divided by five (C5, CIO. Cl5, C20, C25, C30 and C40), although also irregular isoprenoids and polyterpenes have been reported. Important members of the isoprenoids a.o. are the carotenoids, gibberellins, abscisic acid, some Cytokinins, sterols, and the terpenoids, consisting of a.o. monoterpenes, sesquiterpenes, diterpenes, triterpenes, tetraterpenes and polyterpenes (rubbers), etc. Most of these compounds occur free but they can also be modified, or derivatized as esters and glycosides, or attached to proteins. Among the isoprenoids there are many compounds with biological activity, for example as plant growth regulator (gibbereUins, abscisic acid, cytokinins). and in the interaction between plants and other organisms (for example as anti-microbials, infochemicals and as the isoprenoid germination stimxilants that are exuded by the roots of some plant species and induce the germination of parasitic weed seeds).
Mono- and sesquiterpenes, the CIO and CIS branch of the isoprenoid family, were investigated for their economically interesting value as flavor and firagrance compounds in foods and cosmetics and their anti-carcinogenic and antimicrobial properties. Mono- and sesquiterpenes have also been shown to be of ecological significance, for instance in the interaction between plants, plants and insects/spider mites and plants and microorganisms. Therefore, plants producing mono- and sesquiterpenes have been investigated by many authors and this has resulted in a better understanding of the biochemical pathways leading to the formation of these compounds and their derivatives.

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Linalool is an acyclic monoterpene alcohol that has a peculiar creamy floral, sweet taste. In Clarkia breweri (Onagraceae) linalool, amongst other compounds, is responsible for the attraction of pollinating moths. Linalool is one of the volatile compounds released as a semiochemical after herbivore attack in some plants and as such may attract predators of the herbivores. The sweet taste of linalool makes it suitable to enhance the blueberry flavor of foodstuffs especially of beverages (US Patent 4041185). Furthermore, linalool is known to have a broad-spectrum antimicrobial activity. It is reported by Pattnaik et al. (Microbios 89: 39-46. 1997) to display antibacterial activity against Gram-positive and Gram-negative bacteria as well as antifungal activity against yeast-like and filamentous fungi.
Nerolidol, the sesquiterpene analogue of the monoterpenoid linalool, is a component of many essential oils and flower headspaces (Bauer et al.. Common Fragrance and Flavor Materials. Preparations, Properties and Uses, VCH Verlaggesellschaft, Weinheim, Germany, 1990; Knudsen et al, Phytochemistry 33: 253-280, 1993). Nerolidol has been reported to have anti-microbial activity. EP 0420630A2 describes the use of nerolidol in an antiplaque oral composition. Bouwmeester et al (Plant Physiol. 121: 173-180. 1999) for cucumber and Lima bean and Degenhardt and Gershenzon (Planta 210: 815-822, 2000) for maize showed that nerolidol biosynthesis is induced upon respectively spider mite or Spodoptera feeding. The enzyme responsible for the formation of nerolidol catalyses the regulatory step in the formation of the important signalling molecule 4,8-dimethyH,3(£),7-nonatriene. Both nerolidol and 4,8-dimethyl-l,3(E),7-nonatriene are important constituents of the volatile blend produced in maize upon feeding of beet army worm larvae (Turlings et al, Science 250:1251-1253.1990; Degenhardt and Gershenzon, 2000) and in gerbera in response to feeding of spider mites (Krips et al., J. Chem Ecol 1999), Also in the headspace of several flowers, nerolidol is an important constituent often together with 4,8-dimethyl-l,3(E),7-nonatriene (Kaiser, In: Perfumes: Art, Science and Technology, Elsevier Science Publishers, Essex, UK, pp 213-250, 1991; Knudsen et al., 1993). Nerolidol has also been reported as a constituent of pheromone mixtures of a number of insects and spider mites (Aldrich-JR; Lusby-WR; Knchansky-JP, Experientia. 1986, 42: 5, 583-585; Regv-S; Cone-WW. Environmental-Entomology

1976, 5: 1, 133-138) and has been described as being miticidal if formulated in a controlled release substrate (US patent 4775534).Also, nerolidol has been reported to be an extremely effective repellent of mosquitoes.
From a number of plants, several cDNAs encoding enzymes involved in the biosynthesis of monoterpenoids have been isolated such as S-hnalool and i?-hnalool synthases (Cseke et al., Mol Biol. Evol. 15: 1491-1498, 1998; Jia et al., Arch Biochem Biophys 372: 143-149, 1999), (-)-4S limonene synthase (Colby et al., J Biol Chem 268: 23016-23024, 1993; Bohlmann et al., J Biol Chem 272: 21784-21792, 1997).
WO 9715584 describes the use of S-linalool synthase, an acyclic monoterpene synthase, in the genetic engineering of scent production. The use of the limonene (monoterpene) cyclase in the control of corn rootworm, by inserting a nucleotide sequence coding for limonene cyclase into the plants is described in WO 9637102. In WO 0022150 the use of a hmonene synthase, Unalool synthase and combination of limonene and carveol synthase (actually called hmonene hydroxylase) for the control of insects is described. However, terpenoid products were only formed in combination with a GPP synthase.
The enzymes involved in the production of precursors for the synthesis of the primary monoterpene skeletons are all active in the plastids, since all cloned genes of this pathway until now have plastid targeting signals. Recently, for one enzyme, (4S)-limonene synthase, localisation in the leucoplasts of the secretory cells in Mentha spicata has been demonstrated with immunogold labeling. The plastid targeting signals indicate that isoprenoid precursors for monoterpene metabolism are formed in the plastids, although some partitioning of these precursors between the different cellular compartments in plants has been shown to occur. Unlike other monoterpene (and diterpene) cyclases that bear cleavable transit peptides of 50-70 amino acids, the S-linalool synthase cDNA isolated by Pichersky and co-workers encodes a protein with an apparent cleavable peptide of maximally only eight amino acids long. Nevertheless, typical plastid targeting signal characteristics were found in the first

60 amino acids of the cDNA, supporting that the linalool synthase enzyme, as expected for a monoterpene synthase, is indeed targeted to the plastids. Two independent cDNA clones encoding 5-epi-aristolochene synthase (EAS) from tobacco have been isolated and characterised by Facchini and Chappell (Proc Natl Acad. ScL USA, 89:11088-11092, 1992). Back and Chappell described the cloning and bacterial expression of vetispiradiene synthase found in Hyoscyamus muticus (J, BioL Chem., 270(13):7375-7381. 1995). The cDNA encoding amorphα-4,ll-diene synthase, an intermediate in the biosynthesis of the anti-malarial artemisinin, was isolated and characterised by Merclce et al. {Arch. Biochem. Biophys., 381(1):173-180, 2000). Sesquiterpene biosynthesis is compartmentalised to the cytosol, and none of the sofar isolated sesquiterpene synthases bear any targeting signal. Farnesyl diphosphate (FPP) is present in every living organism and it is the precm'sor of a large number of primary and secondary metabolites. It has been established that FPP is the precursor of all sesquiterpenoids. There are several thousands of different sesquiterpenoid compounds identified in many Uving organisms. Examples are the bitter sesquiterpene lactones such as sonchuside A and C, and cichorilide A in chicory (De Kraker et aL, Plant Physiol 117: 1381-1392, 1998). The first committed step Ln the biosynthesis of these compounds is catalysed by a germacrene A synthase which was cloned from chicory (PCT/EP 0002130). Other examples are the cloning of three sesquiterpene synthases ((E)-α-bisabolene, 5-selinene , and γ-humulene synthase) from grand fir (WO 99/37139; Bohlmann et al., proc Natl Acad Sci, USA, 95: 6756-6761), and a germacrene C synthase from tomato (Colby et al., Proc Natl Acad Sci, USA 95: 2216-2221). The use of the amorphα-4,ll-diene synthase in the engineering of artemisinin biosynthesis is described in EP 0 982 404 Al. However, the putative sesquiterpene synthase responsible for the formation of the biologically important nerolidol has never been cloned.
The use of recombinant DNA technology to introduce resistance based on secondary metabolites in plants has had only limited success. For example, Hain et al (Nature 361, 153-156, 1993) succeeded in introducing fungal resistance in a number of plant species by the introduction of the resveratrol synthase cDNA, that they isolated from grape. Although there are reports on the anti-microbial and insecticidal effects of

specific terpenoids, resistance against fungi as a result of the expression of a terpene synthase in plants has not been reported sofar.
The invention provides an isolated or recombinant nucleic acid or functional fragment thereof encoding a proteinaceous molecule essentially capable of isoprenoid bioactive compound (herein also identified as flavor, fragrance and/or bio-control agent) synthesis when provided with a suitable substrate under appropriate reaction conditions. Presently, the main way to produce plant flavor (for ease of reference with flavor also fragrances are generally meant) compounds is by the synthetic route. Synthetic organic chemicals constitute more than 80-90% (by weight and value) of the raw materials used in flavor and frragrance formulations. However, problems often exist concerning production. Extaction from intact plants and conventional fermentation are currently providing alternative routes for the commercial production of flavor/ aroma chemicals. However, the demand for natural flavors by the consumer has been steadily increasing, and demand often outstrips supply. In many cases sought after flavor compounds can not easily be isolated. An understanding of the precursors and characterization of genes encoding enzymes involved in diverse pathways leading to the formation of flavors is essential for the production of natural flavors. The nucleic acids and their encoded proteinaceous molectdes of the present invention are involved in the biosynthetic pathway for terperoid production and as such they provide new means and methods for the in-vivo and in-vitro biotechnological production of bio-flavours, natural flavor chemicals and bio-control compounds.
In addition the nucleic acids and their encoded proteinaceous molectdes of the present invention and products synthesized are essentially capable acting as potent bio-control agents alone or in combination.
Fungi and bacteria have become an increasing threat to humans. Opportmistic microbial infections have increased dramatically in the last two decades and have become a significant cause of morbidity and mortality. Over recent years, the firequency of life-threatening fungal infections has increased dramatically, making fungal infections now responsible for nearly 40% of all deaths from hospital-acquired infections. Increased numbers of patients with an impaired immune system (such as

due to ageing, severe bums, AIDS, chemotherapy against cancer, or immunosuppressive therapy for organ transplants), together with a growing list of potential pathogenic fungi and bacteria are recognized as factors contributing to tliis rising public health-hazard. There is only a limited set of bio-control compounds available, and resistance to existing bio-control drugs is becoming a problem of increasing concern. Also clinically used antimycotics may show harmful side effects.
Fungi are responsible for substantial economic losses due to food spoilage caused by highly dangerous toxins (mycotoxins). To add to this problem food additives to prevent fungal contamination may also be potentially carcinogenic. Additionally plant pathogenic micro-organisms cause huge crop losses and this has promoted the extensive use of pesticides all over the world. Some pesticides have deleterious effects on other organisms than the pests they are intended to control, on water quality, and on the environment in general. Current antimicrobials are often not specific enough, and several microbial species exhibit increasing resistance to these pesticides. There is a need to develop new and more advanced bio-control agents with novel modes of action and broad spectra directed against plant and animal pathogens. The nucleic acids and their encoded proteinaceous molecules of the present invention involved in terpenoid biosynthesis, as such provide a new method for the in-vivo and in-uitro biotechnological production of natural and more specific anti-microbials or bio-control agents, for example antifungals.
The nucleic acid as used herein refers to an oligonucleotide, nucleotide or polynucleotide, and fragments or portions thereof and to DNA or RNA of genomic or synthetic origin which maybe single- or double-stranded, and represents the sense or antisense strand. A proteinaceous molecule as used herein refers to a molecule comprising peptide or protein. Natural flavor synthesis as used herein refers to flavor and fragrance compounds synthesized that are identical to their natural counterpsurts. Natural counterpart as used herein refers to products that are obtained directly from plants and sometimes from animal sources by physical procedm^es. Sjrnthetic flavors refers to nature identical compoimds that are produced synthetically but are chemically identical to their natural counterpart. Nature-identical compounds are with few exceptions the only synthetic compounds used in flavors in addition to natural products. Artificial flavor synthesis refers to flavor

compounds that have not been identified in plant or animal products for human consumption. The nucleic acids of the present invention pave the way for the production of artificial flavors using techniques known in the art such as for example combinatorial biosynthesis, metabolic pathway engineering, gene shuffling, directed evolution of proteins etc. Bio-control synthesis as used herein refers to bio-control compounds synthesized which can act as an bio-control agent. A bio-control agent as used herein refers to a compound, which can at least in part suppress or inhibit or restrict the growth of a pathogenic organism (e.g. fungi, bacteria etc.), that is a compound that has anti-pathogenic activity.
The invention further provides for a nucleic acid or functional fragment thereof wherein said nucleic acid encodes a proteinaceous molecule essentially capable of synthesizing at least a monoterpene alcohol linalool when contacted with geranyl diphosphate (GPP) and/or at least a sesquiterpene alcohol nerolidol when contacted with farnesyl diphosphate (FPP) under appropriate reaction conditions. The definition 'functional fragment thereof means that a particidar subject sequence may vary firom the reference sequence by one or more substitutions, deletions, or additions, the net effect of which does not result in an adverse functional dissimilarity between the reference and the subject sequence. It may be advantageous to produce a nucleic acid according to the invention or derivatives thereof possessing a substantially different codon usage. It is known by those skiUed in the art that as a result of degeneracy of the genetic code, a multitude of gene sequences, some bearing minimal homology to the nucleotide sequences of any known and any naturally occurring genes may be produced. The invention includes possible variation of the nucleic add sequence that could be made by selecting combinations based on possible codon choices. In addition deliberate amino acid substitution may be made on the basis of similarity in polarity, charge, solubility, hydrophobicity, and/or the amphipathetic nature of the residues as long as the biological activity of the polypeptide is retained. In a preferred embodiment said nucleic acid encodes a terpene cyclase which has a combined nerolidol and linalool synthesizing capacity. Nerolidol is a sesquiterpene analog of the monoterpenoid linalool. The enzymes involved in the production of precursors for the Synthesis of the primary monoterpene skeletons have been shown to be active in the plastids. The ability of said terpene cyclase to synthesize linalool

appears to be influenced by the presence of a plastid targeting signal sequence that is rich in hydroxylated and basic residues. Sesquiterpene biosynthesis is compartmentalised to the cytosol, and none of the sofar isolated sesquiterpene synthases bear any targeting signal. However, the present invention shows that monoterpenes can also be produced by cytosolic monoterpene synthases. Apparently the substrate GPP is present in the cytosol. The invention shows that the production of sesquiterpenes in the cytosol is hampered by a lack of substrate. The co-expression of a cytosolic FPP-synthase or the transformation with a fusion construct of sesquiterpene synthase and FPP synthase is now provided to overcome this problem. An additional solution is the targeting of sesquiterpene biosynthesis to other cell compartments by adding or changing a targeting signal to/of the sesquiterpene synthase and/or co-transformation of an FPP synthase with the same targeting or transformation with a targeted fusion construct of sesquiterpene synthase and FPP synthase. In addition to FPP synthase, other enzymes catalyzing committed steps in the biosynthesis of GPP and FPP through the mevalonate and non-mevalonate pathway can be coupled to or co-expressed with monoterpene and sesquiterpene synthases to increase the levels of monoterpenes and/or sesquiterpenes produced. These enzymes can be directed (by adding, changing and removing targeting signals) to different compartments (i.e. mitochondria, chloroplasts, chromoplasts, leucoplasts, peroxisomes (see also example 7).
The invention thus provides a nucleic acid according to the invention encoding a proteinaceous molecule provided with a targeting signal, such as a plastid targeting or a mitochondrial targeting signal, or a targeting signal to any other organel or a nucleic acid according to the invention encoding a proteinaceous molecule without such signal, depending on where synthesis is req\ured. The invention thus provides a nucleic acid according to the invention encoding a proteinaceous molecule essentially capable of isoprenoid bio-active compound synthesis in the cytosol in a cell when provided with a suitable substrate under appropriate reaction conditions. Similarly, it provides a nucleic acid according to the invention encoding a proteinaceous molecule essentially capable of-isoprenoid bio-active compound synthesis in a plastid in a cell or in a mitochondrium in a cell when provided with a suitable substrate imder appropriate reaction conditions.

In a preferred embodiment said nucleic acid as provided herein is provided with a nucleic acid encoding a targeting signal and/or remnants of a targeting signal. Preferably said targeting signal is a plastid targeting signal. Said plastid targeting signal is preferably located in the N terminus (N-terminal transit peptide) and may have a high abundance of serine residues and/or theronine and/or a low number of acidic residues and/or rich in hydroxy la ted and basic residues. In one preferred embodiment said targeting signal has a F (Phe), K (Lys), V (Val), F (Phe), N (Asn) motif and/or a D (asp) S (Ser), L (Leu), L (Leu), Xaa, S (Ser), S (Ser) motif, where Xaa is preferably P(pro) or S (Ser). In another, the target signal RRxxxxxxxxW is preferred. In particvdar the invention provides a nucleic acid encoding an essentially sesquiterpene synthase bioactive fragment, said nucleic acid provided with a targeting signal to provide the encoded gene product with monoterpene Synthase activity, or a nucleic acid encoding an essentially monoterpene synthase bioactive fragment, said nucleic acid deprived from an essentially plastid targeting signal to provide the encoded gene product with sesquiterpene synthase activity, and thus provides the various enzymes with a different activity as would be expected.
It is understood that through convergent or divergent evolution new proteins with altered functions may be created by this route. The mutations that lead to divergence are mostly single base substitutions that engender individual amino acid replacements, although other events leading to deletions or insertions also occur. The mutations may be in a nucleic acid comprising the transit peptide and/or the open reading frame (ORF). The new protein usually contains many of the pre-existing features. The original biological function may be restored by reversing mutations (e.g. single base substitutions) using techniques known in the art (e.g. site directed mutagenesis).
In a preferred embodiment through a single base substitution in a predecessor sequence of said nucleic acid sequence (e.g. H64NORL) the N-terminal transit peptide is restored- Restored as used herein means that a stop codon in the target signal is removed, for example through a single base substitution, so that translation begins at the first ATG (Met) upstream of the target signal/transit sequence or target signal remnant. The predecessor sequence of said nucleic sequence is a sequence (common ancestor sequence) which has a stop codon in the target signal or the target

signal remnant so that the translation of the protein begins at a second ATG (Met) truncating the target signal or the target signal remnant. It is conjectured that the presence of the restored target signal or target signal remnant influences the synthesis of hnalool and/or nerolidol.
The invention provides for a nucleic acid according to the invention wherein said proteinaceous molecule comprises a terpene synthase/cyclase. Preferably said proteinaceous molecule comprises a terpene synthase (cyclase), the properties of which should resemble those of other terpene synthases (cyclases). The invention further provides a nucleic acid according to the invention wherein said proteinaceous molecule comprises a sesquiterpenoid synthase/cyclase. Sesquiterpenoid synthases/cyclases participate in the biosynthesis of most sesquiterpenoids. Ionization of FPP to the farnesyl cation is the first step in the biosynthesis of a large number of sesquiterpenes. The products of many of the sesquiterpenoid synthases/qrclases catalyzing the formation of a terpenoid skeleton from the respective diphosphate substrates (FPP) are mostly cyclic hydrodrocarbons, with a few exceptions such as for example the acyclic sesquiterpene alcohol nerolidol. None of the sofar isolated sesquiterpene synthases bear any targeting signal.
The invention further comprises a nucleic acid according to the invention wherein said proteinaceous molecule comprises a nerolidol synthase/cyclase protein or functional fragment thereof. The nerolidol synthase/cyclase protein is essentially capable of the synthesis of the acyclic sesquiterpene alcohol nerolidol.
The invention provides a nucleic acid wherein said nerolidol synthase/cyclase comprises (3S)-(E)-nerolidol synthase. The inventioq. furtiier comprises a nucleic acdd according to the invention wherein said sesquiterpene alcohol nerolidol comprises trans-nerolidol. The invention further comprises a nucleic acid according to the invention wherein said monoterpene alcohol linalool comprises S-linalool.
The invention provides for a nucleic add according to the invention wherein said nucleic acid encodes a proteinaceous molecule comprising an amino acid sequence or functional fragment thereof that is at least 50% identical to H64MUT sequence, more preferred 53 or 60% homologous, and even more preferred 70, 80, 90, 95 or 99% homologous to the sequence as shown in Figure 2 or functional fragment thereof

Homology is generally over the full-length of the relevant sequence shown herein- As is well-understood, homology at the amino acid level is generally in terms of amino acid similarity or identity. Similarity allows for "conservative variation", i. e. substitution of one hydrophobic residue such as isoleucine, valine, leucine or methionine for another, or the substitution of one polar residue for another, such as arginine for lysine, glutamic for aspartic acid, or glutamine for asparagine. Deliberate amino acid substitution may be made on the basis of similarity in polarity, charge, solubility, hydrophobicity, and/or the amphipathetic nature of the residues as long as the biological activity of the polypeptide is retained. In a preferx'ed embodiment, all percentage homologies referred to herein refer to percentage sequence identity, e.g. percent (%) amino acid sequence identity with respect to a particular reference sequence can be the percentage of amino acid residues in a candidate sequence that are identical with the amino acid residues in the reference sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, without considering any conservative substitutions as part of the sequence identity. Amino acid similarity or identity can be determined by genetic programs known in the art.
The invention further provides a nucleic acid or functional fragment thereof according to the invention wherein said nucleic acid encodes a proteinaceous molecule essentially capable of the synthesis of at least one monoterpenoid when contacted with geranyl diphosphate (GPP) under appropriate reaction conditions. The invention further provides a nucleic add according to the invention wherein said nucleic acid encodes a proteinaceous molecule essentially capable of the synthesis of at least one monoterpenoid, wherein said monoterpenoid comprises α-pinene and/or β-pinene (bicyclic terpene hydrocarbons) and/or sabinene and/or β -myrene (acyclic monoterpene) and/or α-phellandrene and/or β-phellandrene and/or α-terpinolene and/or α-terpineol and/or γ-terpinene. Preferably said proteinaceous molecule comprises a terpene synthase (cyclase), the properties of which should resemble those of other terpene synthases (cyclase). Even more preferred said proteinaceous molecide comprises a monoterpenene synthase/cyclase. Preferably said monoterpenoid comprises an olefinic monoterpenoid.

The invention further comprises a nucleic acid according to the invention wherein said nucleic acid encodes a proteinaceous molecule comprising an amino acid sequence or functional fragment thereof that is at least 50% identical to SOSV sequence (see figure 6), more preferred 53 or 60% homologous, and even more preferred 70, 80 or 90 % homologous to the sequence as shown in figure 6 or functional fragment thereof, Preferably said nucleic acid does not contain an insertion of two cytosine residues causing a frame-shift followed by a stop codon giving rise to a truncated open reading frame (ORF), as depicted in figure 6 and 7.
The invention further comprises a nucleic acid encoding a proteinaceous molecule according to the invention obtainable from a eukaryote. A eukaroyte as used herein comprises a cell or organism with a membrane-bound, structurally discrete nucleus and other well-developed subcellular compartments. Eukaryotes as used herein include all organisms except viruses, bacteria, and cyanobacteria (blue-green algae). Preferably said nucleic acid is obtainable from strawberry and/or maize and/or tea and/or cucumber and/or lima bean and/or cotton and/or thyme species and/or citrus species and/or eucalypt species and/or grapefruit and/or fungi and/or yeasts.
The invention further comprises a nucleic acid encoding a proteinaceous molecule according to the invention obtainable from a prokarote. A prokaryote as used herein comprises a cell or organism lacking a membrane- bound, structurally discrete nucleus and other subcellular compartments e.g. bacteria, including archaebacteria and cyanobacteria (blue green algae).
The invention further comprises a nucleic acid encoding a proteinaceous molecule according to the invention obtainable from invertebrate animals. An arthropod is a member of a phylum of invertebrate animals that includes insects, arachnids (spiders and mites e.g. spider mites (Tetranychus urticae), aphids (e.g. Aphis gossypii, Myzus persicae), and thrips (Frankliniella occidentalis) and crustaceans (crabs, lobsters, pillbugs, shrimp, etc.).
In a preferred embodiment said nucleic acid encoding a proteinaceous molecule according to the invention is obtainable from strawberry. The invention further provides a nucleic acid according to the invention wherein said nucleic acid expression is repressed by auxin during fruit maturation- Indole-3-acetic acid or

auxin is a plant hormone that plays key roles in regulating cell division, extension, and differentiation.
The invention provides a proteinaceous molecule encoded by a nucleic acid according to the invention. The invention further provides a vector comprising a nucleic acid according to the invention. Preferably said vector is a recombinant expression vector comprising a coding sequence which is operably linked to a promoter sequence capable of directing expression of said coding sequence in a host cell for said vector, and a transcription termination sequence, in which the coding sequence is a nucleic acid according to the invention. Preferably said nucleic acid has been provided with means for nuclear targeting and/or integration in a host genome.
Methods which are well known in the art can be used to construct expression vectors containing the nucleic acid of the invention, and appropriate transcriptional and translational controls. These methods include in-vitro recombinant techniques. Exogenous transcriptional elements and initation codons can be used and also can be of various origins, both natural and synthetic. The efficiency of expression may be enhanced by the inclusion of enhancers appropriate to the cell system in use. In the case of plant expression vectors, the expression of a nucleic acid of the invention may be driven by a number of previously defined and yet to be defined promoters, including inducible and developmentally regulated promoters. The invention further contemplates the use of the individual promoters of the nudeic add of the present invention for this purpose. In particular any promoters particularly responsive to ripening events, wound-inducible or specific inducible promoters (e.g. spider mite, insect etc. inducible promoters, which can be isolated from plants that were fed upon by for example spider mites oir insects), may be used to drive the tissue specific expression of said nudeic add. In addition, viral promoters such as the 35S and the 19S promoters of CaMV may be used alone or in combination with the omega leader sequence firom TMV. Promoters or enhancers derived firom the genomes of plant cells, tissue specific promoters i.e firuit specific promoters, Fbp7 (Columbo et aL 1997; Plant Cell 9; 703-715), 2AH promoter (Pear et cd., 1989, Plant Molecular Biology, 13:639-651), small subimit of Rubisco (Corruzzi et al., 1984; EMBO J 3:16; Broglie et al., 1984 Science 224:838-843) or timing specific promoters such as ripening specific promoters (the E8 promoter, Diekman and Fisher, 1988,

EMBO J, 7:3315-3320) may be lised. Sliitable terminator seqliences incllide that of the Agrobacterilim tlimefaciens nopahne synthase gene (Nos 3' end), the tobacco riblilose bisphosphate carboxylase small sliblinit termination region; and other 3' regions known in the art. Methods known in the art can be lised to constrlict recombinant vectors which will express 'sense' or 'antisense' nlicleic acid. Antisensa or partial sense or other techniqlies may also be lised to redlice the expression of said nlicleic acid leading to the prodliction of a flavolir, fragrance and/or bio-control compolind. Flill length sense techniqlies may be lised to increase or redlice the expression of said nlicleic acid leading to the prodliction of a flavor and bio-control compolind.
The invention flirther provides a replicative cloning vector comprising a nlicleic acid according to the invention and a replicon operative in a host cell for said vector. The invention contemplates the lise of yet non-described biological and non biological based expression systems and novel host(s) systems that can be can be litilized to contain and express the nlicleic acid of the invention. The definition host cell as lised herein refers to a cell in which an foreign process is execlited by bio-interaction, irrespective of the cell belongs to a linicelllilar, mlilticelllilar, a differentiated organism or to an artificial cell, cell cliltlire or protoplast.
The invention flirther provides a host comprising a nlicleic acid according to the invention or a vector according to the invention. A variety of vector/host expression systems can be litilized to contain and express the nlicleic acid of the invention. These incllide micro-organisms slich as bacteria (e.g. E coli, B slibtilis, Streptomyces, Pselidomonads) transformed with recombinant bacteriophage, plasmid or cosmid DNA expression systems, yeast (eg, S. cerevisiaei, Klliyvefomyces lixctis, Pichia pastoris, Hansenlila polytnorpha, Schizosacch. Pombe, Yarrowia) transformed with yeast expression vectors; filamentolis fungi (Aspergilllis nidlilaiis, Aspergilllis orizae, Aspergilllis niger) transformed with filamentolis flingi expression vectors, insect celi systems transfected with virlis expression vectors (eg baclilovirlis, adenovirlis, herpes or vaccinia virlises); plant cell systems transfected with virlis expression vectors (e.g. caliliflower mosaic virlis, CaMV, tobacco mosaic virlis, TMV) or transformed with bacterial expression vectors (e.g Ti or Pbr322 plasmid); or mammalian ceE systems (Chinese hamster ovary (CHO), baby hamster kidney (BHK), Hybridoma's, inclliding

cell lines of molise, monkey, hliman and the Like. A host strain may be chosen for its ability to modlilate the expression of the nlicleic acid or to process the expressed proteinaceolis moleclile in the desired fashion. Slich modifications of said proteinaceolis moleclile incllide acylation, carboxylation, glycosylation, phosphorylation and hpidation. Post translation processing which cleaves a 'prepro' form of said proteinaceolis moleclie may also be important for correct insertion, folding and/or flinction. Different host cells which have the correct celllilar machinery and characteristic mechanisms for slich post-translational activities maybe chosen to enslire correct modification and processing of the introdliced, foreign proteinaceolis molecule.
The invention flirther provides a host comprising a nlicleic acid according to the invention or a vector according to the invention wherein said host comprises a prokaroytic cell. The invention flirther provides a host comprising a nlicleic acid according to the invention or a vector according to the invention wherein said host comprises a elikaryotic cell.
The invention further provides a host comprising a nlicleic acid according to the invention or a vector according to the invention wherein said host comprises a plant • and propagating material thereof. The invention is particlilarly liseflil for enabling plants to prodlice linalool, nerolidol or a combination of the two. This enables breeding of plants with improved flavor/fragrance as described for linalool alone in WO 9715584, or improved resistance against micro-organisms or insects as described in Examples 8, 9, 12, 13 and 14 and WO 0022150 for linalool that had however to be co-expressed with a GPP synthase.
The bacterial diseases to which resistance is provided herein incllide blit are not limited to:
Erwinia spp. (e.g. E. amylovora (fire blight) and E. carotovora), Clavibacter spp. (e.g. C. michiganense pv. Sepedoniclim (bacterial ringspot potato), Corynebacterilim spp., Pselidomonas spp. (e.g. P. syringae pv. tomato), Xanthomonas spp. (X. campestris and X. vesicatoria), and Agrobacterilini spp.
The flingal diseases to which resistance is provided herein incllide blit are not limited to:

Powdery mildew flingi (Sphaerotheca spp. (e.g. S. pannosa var. rosa, (rose), S. hlimiili (hop), S. fliliginea (clictixbits)), Podosphaera lelicotricha fapple), lincinlila necator Cgrape
0, Erysiphe spp,(e.g. E. cichoracearlim (cliclirbits, tomatoj, E, polygoni (beet)), LeDeillida iaxirica (tomato), Microsp/iaera elionymi (sqliash)), Botrytis spp. (e.g. B. cinerea (grey mold)), Cladosporilim spp, (e.g. C. flilvlim. (in tomato)), Sphaeropsis spp, (e.g. Sphaeropsis sapinea (tip bhght of pine), Cercospora spp. (C. beticola in beet, C. zeae-maydis in com, C. sorghi in sorghlim), Flisarilini spp. (e.g. F. oxysporlim f. nivelim (wilt on watermelon) F. graminearlim and F. moniliforme (scab on wheat) F. moniliforme, F. oxysporlim, F. slibgllitinans, F. proliferatlim), anthracnose diseases (Apiognomonia veneta (in Sycamore, ash, oak, maple, and walnlit), Colletotrichlim trifolii (Alfalfa anthracnose), Colletotrichlim coccodes (black dot in potato)), rlist diseases (e.g, Pliccinia recondita (leaf rlist in wheat) and liromyces appendiclilatlis (rlist in bean)), Phytophtora spp. (P. infestans (late blight on potato), P. sojae (blight on soybean), P. megasperma t sp. medicaginis (root rot in alfalfa)), spoilage flingi {Gibberella spp., Diplodia spp., Penicillilim, Aspergilllis spp. Penicillilim spp,, Peacilomyces spp.), Verticillilim spp. (e.g. V. albo-atrlim (black root rot in strawberry), Sepioria spp. (e.g. S. tritici and S. avenae f. sp. triticea (Septoria in wheat), S. lycopersici (Septoria leaf spot in tomato)), Sclerotinia spp. (e.g. S. sclerotiorlim (white mold of beans), Aphanomyces spp. (e.g,A cochlioides (root rot in sligar beet), Altemaria spp. (e.g. A. solani (early blight in tomato), Magnaporthe spp. (e.g. M. grisea (blast in rice)) Insects
The insects to which resistance is provided herein incllide blit are not limited to Lepidoptera, Orthoptera, Homoptera, Hemiptera, especially sqliash bligs {Anasa tristis); green stink blig {Acrosternlim. hilare); Riptortlis clavatlis; Coleoptera, especially, Colorado potato beetle (Leptinotxxrsa decemlifieata); three-lined potato beetle (Lema trilineclia); asparaglis beetle (Crioceris asparagi); Mexican bean beetle {Epilachna varivestis); red flolir beetle (Tribolilim castanelim); conflised flolir beetle (Tribolilim conflislim); the flea beetles (Chaetocnema spp., Haltica spp. and Epitrix spp.)] com rootworm (Diabrotica Spp.); cowpea weevil (Callosobrlichlis maclilatlis); boll weevil (^thonomlis grandis); rice weevil (Sitophillis oryza); maize weevil (Sitophlilis zeamais)] granary weevil (Sitophillis granarilis); Egyptian alfalfa weevil

(Hypera postica)\ bean weevil {Acanthoscelides obtectlis); lesser grain borer {Rhyzopertha doniinica); yellow meal worm (Tenebrio molitor); Thysanoptera, especially, western flower thrips {Frankliniella occidentalis); Diptera, especially, leafminer spp. (Liriomyza trifolii); plant parasitic nematodes especially the potato cyst nematodes (Globodera spp.), the beet cyst nematode (Heterodera schachtii) and root knot nematodes (Meloidogyne spp.)-
Resistance can de determined by performing the appropriate test with the particlilar organism blit can be predicted as well by determining terpene content slich as demonstrated in figlire 30 and example 13 herein.Plant as lised herein refers to elikaryotic, alitotrophic organisms. They are characterized by direct lisage of solar energy for their primary metabolism, their permanent cell wall and in case of mlilticelllilar individlials their open linlimited growth. In case of heterotrophic plants, the organisms are in an evollitionary context essentially derived from alitotrophic plants in their strlictlire and metabolism. The invention provides a plant or a part, slich as a stem, leave, tliber, root, frliit or seed or propagating material thereof transformed with the expression vector according to the invention. The invention flirther provides a plant or part thereof which contains within its genome a vector according the invention.
The invention provides a host comprising a nlicleic acid according to the invention or a vector according to the invention wherein said host comprises a plant celLl 'Plant Cell' as lised herein is any self-propagating cell bolinded by a semi permeable membrane and containing one or more plastids. Slich a cell reqliires a cell wall if flirther propagation is required. Plant celi as lised herein may be part of a whole plant or may be an isolated cell or part of a tisslie which may be regenerated into a whole plant and incllides for example, seeds, slispension cliltlires, embryos, meristematic regions, callolis tisslies, protoplasts, leaves, roots, shoots, gametophytes, sporophjrtes, pollen and microspores. The vector constrlicts according to the invention can be introdliced into plant cells by direct DNA transformation, or pathogen mediated transfection. The procedlire or method for preparing a transformant can be performed according to the conventional techniqlie lised in the fields of moleclilar biology, biotechnology and genetic engineering. Maniplilation of nlicleic add in plant cells may be carried olit lising the Cre/lox site specific

recombination system as olitlined in patent application W09109957. The target plant may be selected from any monocotyledonolis or dicotyledonolis plant species. Exemplary plants incllide potato, tomato, petlinia, maize, sorghlim, cotton, soybean, beans, rape, alfalfa, asparaglis, sweet potato and chrysanthemlim. However, it is not to be constrlied as limiting, in as mlich as microbes and insects may infest many other crops. Thlis, the methods of the invention are readily applicable to nlimerolis plant species, if they are folind to be slisceptible to the microbes or insect species listed hereinabove, inclliding witholit limitation, species from the genera Medicago, Tf'ifolilini, Vigna, Citrlis, Daliclis, Arabidopsis, Brassica, Raphanlis, Sinapis, Capsiclim, Lycopersicon, Nicotiana, Solarilim., Helianthlis, Bromlis, Asparaglis, Paniclim, Pennisetlini, Cliclimis, Glycine, Lolilini, Triticlim and Zea.
The invention flirther provides a host comprising a nlicleic acid according to the invention or a vector according to the invention wherein said host comprises an insect cell. Insect cells slich as silkworm cells or larvae themselves may be lised as a host. For example in one slich system, Alitographa califoraica nliclear polyhedrosis virlis (AcNP\0 is lised as a vector to express foreign nlicleic acid in Spodoptera frligiperda cells or in Trichopllisia larvae. The nlicleic acid of the invention may be cloned into the nonessential region of the virlis, slich as the polyhedrin gene, and placed linder control of a polyhedrin promoter. Sliccessflil insertion of the nlicleic acid will render the polyhedrin gene inactive and prodlice recombinant virlis lacking coat protein coat. The recombinant virlises are then lised to infect S frligiperda cells or Trichopllisia larvae in which the nlicleic acid is expressed [Smith et al. (1993) J Virol 46:584; Engelhard et oL, (1994) Proc Naliacad Sci, 91: 3224-7].
The invention flirther provides a host w^herein said vector according to the invention and said host expresses a nerolidol synthase/cyclase protein or polypeptide. Preferably said host exhibits sliitable glycosyltransferase activity, whereby the prodliced linalool and nerolidol is converted and acclimlilated or stored in said host as its respective linaloylglycoside and nerolidylglycoside. Preferably said host contains the appropriate (indlicible) glycosidase enzyme sliitable for the release of the respective linalool and nerolidol. Alternatively said host is provided with a nlicleic acid coding for a sliitable (indlicible) glycosidase enzyme. Host which contain a nlicleic acid encoding a proteinaceolis moleclile according to the invention may be

identified by a variety of procedlires known in the art. These procedlires incllide, blit are not Limited to DNA-DNA, DNA
-RNA hybridisation, amplification lising probes (portions or fragments of said nlicleic acid), protein bioassay or immlinoassay techniqlies which incllide membrane, sollition or chip based technologies for the detection and/or qliantification of said nlicleic acid and encoded proteinaceolis moleclile.
The invention flirther provides a host wherein said vector according to the invention and said host expresses a monoterpenene synthase/cyclase protein or
polypeptide.
The invention provides a method for prodlicing a flavor, fragrance and/or bio-control compolind comprising a) transforming or transfecting a sliitable host with at least one nlicleic acid encoding a proteinaceolis moleclile according to the invention b) expressing said nlicleic acid in the presence of a sliitable slibstrate c) optionally isolating the formed prodlict. In a preferred embodiment said nlicleic acid incllides a restored target signal or a target signal remnant, i.e. in those case where plastid targeting is reqliired. In a preferred embodiment of the invention is a method to prodlice nerolidol and /or linalool and/or α-pinene and/or β-pinene (bicyclic terpene hydrocarbons) and/or sabinene and/or β-myrcene (acyclic monoteipene) and/or α-phellandrene and/or β-phellandrene and/or α-terpinolene and/or α-terpineol and/or γ-terpinene or mixtlires thereof by a) transforming/transfecting a sliitable host b) expressing at least one nlicleic acid of the invention in the presence of a sliitable slibstrate and c) isolating the formed prodlicts. In a preferred embodiment said host exhibits sliitable glycosyltransferase activity, whereby the prodliced linalool and/or nerolidol is converted and acclimlilated or stored in said host as its respectivie linaloylglycoside and nerolidylglycoside. It is most easy when said host already contains the appropriate (indlicible) glycosidase enzyme siiitable for the release of the respective linalool and nerolidol. This is however not reqliired, expression witholit said glycosyltransferase and/or glycosidase activity is perfectly well sliitable for most plirposes and alternatively said host may even be provided with a nlicleic acid coding for a sliitable glycosidase enzyme, when deemed reqliired. For bio-control acitivity, it is even provided to express the compolinds according to the invention witholit said glycosyltransferase and/or glycosidase activity, and let the bio-control activity partly

depend on said activity in the target organism, e.g. after liptake by an insect the insects saliva, or on the indliction of said activity after herbivory or flingal infection.
A method for prodlicing a compolind according to the invention is provided comprising a) transforming or transfecting a sliitable host with at least one nlicleic acid encoding a proteinaceolis moleclile according to the invention b) expressing said nlicleic acid in the presence of a sliitable slibstrate c) optionally isolating the formed prodlict, wherein said host comprises a microorganism, plant cell or plant. Microorganism as lised herein refers to microscopic organisms slich as for example Archaea, Bacteria, Cyanobacteria, Microalgae, Flingi, Yeast, Virlises, Protozoa, Rotifers, Nematodes, Micro-Crlistaceans, Micro-Mollliscs, Micro-Shellfish, Micro-insects etc.
The invention provides a method for prodlicing a flavor, fragrance and or bio-control compolind in a cell-free lysate expression system comprising expressing at least one nlicleic acid encoding a proteinaceolis moleclile according to the invention in the presence of a sliitable slibstrate and optionally isolating the formed prodlict, wherein said free lysate system contains all the components necessary for expression and processing. Cell-free lysate expression system as lised herein refer to cell-free translation/translocation systems known in the art, slich as for example rabbit reticlilocyte lysate translation system.
The invention flirther provides a flavor and/or bio-control compolind obtainable by a method according to the invention. Preferably said flavor and/or bio-control compolind comprises at least a nerolidol and/or linalool and/or α-pinene and/or 6-pinelie (bicyclic terpene hydrocarbons) and/or sabinene and/or β-myrcene (acyclic monoterpene) and/or α-phellandrene and/or β-phellandrene and/or α-terpinolene and/or α-terpineol and/or γ-terpinene or mixtlires thereof.
The invention flirther provides lise of a flavor compolind according to the invention in the processed food indlistry as an additive. Preferably as a food additive to enhance the flavor of syrlips, ice-creams, ices, frozen desserts, yoglirts, pastries, salices, sweets, confectionery, baked goods etc., and like prodlicts, for example the enhancement of bllieberry flavor (liS Patent 4041185). Strawberry is a poplilar frliit for nattiral flavor ingredients becalise of its flavor, fragrance, aroma and scent. The invention provides the lise of the nlicleic acid according to the invention, for the

indlistrial prodliction of 'fruit' flavors which are natliral to match the odor fidelity of the natliral fruiit. The invention provides for the prodliction of novel flavors, fragrances and/or bio-control agents by the lise of the nlicleic acid according to the invention, alone or in combination, to provide novel avenlies for prodliction. For example, the natliral or the stereochemicaily plire nerolidol may be lised as a slibstrate for the semi-synthesis of flavor and fragrance compolinds or insect repellents as described in liS Patent 005196200A), The compolinds of the present invention may be lised to replace potentially carcinogenic synthetic food additives clirrently lised. The invention provides lise of a flavor and/or bio-control compolind according to the invention as a disinfectant additive for example to obtain natlii*al formlilations and compositions slich as antiplaqlie oral compositions as described in EP 0420630). The invention flirther provides lise of a flavor and/or bio-control compolind according to the invention as a degreasing solvent and/or plasticiser and/or dye carrier.
The invention flirther provides lise of a flavor and/or bio-control compolind according to the invention as a flavoring and/or bio-control agent for oral medications and vitamins. The invention flirther provides lise of a flavor compolind according to the invention for providing additional flavor/aroma in beverages, inclliding alcoholic and non-alcoholic beverages.
The invention flirther provides lise of a flavor compolind according to the invention for enhancing or redlicing plant flavor/aroma/fragrance/scent. The invention flirther provides lise of a flavor compolind according to the invention for enhancing the flavor/aroma of natliral prodlicts and/or synthetic prodlicts and/or artificial prodlicts. The invention flirther provides lise of a flavor compolind according to the invention for the indlistrial synthesis of natlire identical flavor/aroma slibstances. In a preferred embodiment said flavor compolind of the present invention is lised for the prodliction of novel combinations of artificial flavor slibstances.
The invention provides lise of a flavor and/or bio-control compolind according to the invention as a pest control agent. Pest as lised herein is a general term for organisms (rats, insects, mites, micro-organisms etc.) which may calise illness or damage or conslime food crops and other materials im.portant to hlimans/animals. The nlicleic acid of the present invention pave the way throligh plant breeding to

prodlice crops at least more capable of controlling or even eliminating detrimental pest infestations by enabling them to prodlice more terpenoid volatiles (plant volatile allelochemicals) to repel the attacking pest and/or to attract natliral pest enemies to the crop. Preferably said terpenoid volatiles comprise nerolidol and/or linalool. The flavor and/or bio-control compolinds of the present invention can be lised as insecticides, insect repellents, insect pheromones, miticides, scabicides, antimicrobial agents, anti-flingals, anti-herbivore feeding agents etc. For example, nerolidol has been reported to be an extremely effective repellent of mosqliitoes. Formlilations containing natliral nerolidol, prodliced according to the present invention, may therefore be lised in mosqliito control.
In a preferred embodiment said compolind according to the invention is lised for control of the a) interaction between plants and insects b) interaction between plants and micro-organisms c) interaction between one plant and another.
The invention provides lise of a flavor and/or bio-control compolind according to the invention as an anti-microbial agent. Anti-microbial agent as lised herein refers to a compolind which can at least in part slippress or inhibit or restrict the growth of a pathogenic organism (e.g. flingi, bacteria, yeast etc.).
Preferably said compolind may be lised together with at least one other compolind having anti-microbial activity to aligment or slipplement said anti-microbial activity (e.g. said compolind can act synergistically with at least one other anti-microbial compolind). The lise of synergistic combinations of anti-microbial agents has many advantages. One slich advantage is that it minimizes the known risk associated with the lise of potentially deleteriolis anti-microbial agents which can be lised in lower dosages to achieve the same effect. It also lowers risks associated with the lise of non specific/non-selective anti-microbial agents, for example as additives in food and non food prodlicts. Preferably said compolind can be lised for crop treatment programs to redlice or eliminate the lise of harmflil pesticides/biocides [e.g. spray treatments]. It can be incorporated into prodlicts as an bio-control agent [e.g. holisehold materials, detergents, food prodlicts etc.] or applied to prodlicts [e.g. as an external coating to leather prodlicts etc.] to redlice risk of spoilage or contamination.
The invention flirther provides lise of a flavor compolind according to the invention for providing flavor/aroma in cosmetics (inc. soap perflimes, perflime

specialties and bases), creams, slin-protectant prodlicts, hair conditioners, cleaning prodlicts, personal care prodlicts, health care prodlicts (inc. ali mammalian health care prodlicts). The invention flirther provides lise of a flavor compolind according to the invention as a lengthening agent and fixative in perflimes or as a slispension aid for alliminilim salts in anti-perspirants pharmaceliticals (e.g. deodorants).
The invention provides lise of a nlicleic acid according to the invention as a moleclilar marker or diagnostic tool. Preferably as a moleclilar marker for flavor formation [for example nerolidol and /or linalool and/or α-pinene and/or β-pinene (bicyclic terpene hydrocarbons) and/or sabinene and/or β-myrcene (acyclic monoterpene) and/or α-pheliandrene and/or β-phellandrene and/or α-terpinolene and/or α-terpineol and/or γ-terpinene prodliction] in plant breeding. Even more preferred as a moleclilar marker for frliit ripening (for example frliit ripening of strawberry and grapeifruit). The nlicleic acid according to the invention can be lised as markers for the selection of crop species, slich as for example maize, cotton, apple, and cliclimber, and any other crops employing a volatile release defense mechanism, with improved prodliction of volatile terpenoids (e.g. a predator attracting flavor (terpenoid) compolind according to the invention) in response to feeding pests.
The invention flirther provides lise of a flavor and/or bio-control compolind according to the invention in the preparation of a composition. Sliitable basis for compositions are known in the art. Preferably said composition comprises at least nerolidol and/or linalool and/or α-pinene and/or β-pinene and/or sabinene and/or 6-myxcene and/or α-phellandrene and/or β-phellandrene and/or α-terpinolene and/or α-terpineol and/or γ-terpinene, or mixtlires thereof.
The invention flirther provides a composition comprising a flavor and/or bio-control compolind according to the invention. Preferably said compositions are antiflingal, miticidal, or pestiddal. For example a miticidal composition is lisefel for controlling spider mite poplilations. Preferably said compositions comprise slow-release formlilations which can be employed for flimigation plirposes. For example flimigation in agricliltlire for the protection of crops against micro-organisms and pests e.g. insects, mites etc. Preferably said composition is in a form that can be administered to a plant, animal (inclliding hliman), food or non-food prodlict, indlistrial prodlict etc.

The invention provides a composition comprising a flavor and/or bio-control compolind according to the invention which is a pharmacelitical. Sliitable pharmacelitical compositions are known and they may be in dosage forms slich as tablets, pills, powders, slispensions, capsliles, slippositories, injection preparations, ointments, eye drops etc. The invention provides a composition comprising a flavor and/or bio-control compolind according to the invention which is a nelitracelitical.
The invention provides for lise of a composition comprising a flavor and/or bio-control compolind according to the invention for aligmenting or enhancing the aroma and/or taste of food or non food prodlicts and/or protection of food or non food prodlicts against flingal contamination and/or pest infestation. For example chewing glims, medicinal prodlicts, detergents, cosmetics, confectionery etc. Preferably said composition will enhance the shelf life/preservation of food and non-food prodlicts (inc. indlistrial prodlicts).
The invention provides for lise of a composition comprising a flavor and/or bio-control compolind according to the invention for the biological control of pests. For example administrating said composition to a plant. Modes of administration can readily be determined by conventional protocols and may take the form of sprays, dissollible pellets etc.
The invention provides for lise of a composition comprising a flavor and/or bio-control compolind according to the invention for the protection of stored prodlicts. For example for the protection of stored prodlicts against micro-organisms, insects and other pests. For example the protection of potatoes, flowerblilbs, onions etc. against Phytophtora spp, Phoma spp, Flisarilim, Botrytis spp and other stored prodlict pathogens.
The invention provides for lise of a composition comprising a flavor and /or bio-control compolind according to the invention for the prevention or treatment of disease. For example for the treatment of dental caries and/or dental plaqlie and/or skin disorders (dermatological formlilations) and/or immlinoslippressive, anti-lelikaemia and anti-retroviral treatment. A preferred embodiment is that said composition is sliitable for hliman conslimption or external application.
The invention provides for a method of treatment of disease comprising administering a composition according to the invention with a carrier to a sliitable

recipient. Preferably said carrier is a pharmacelitically acceptable carrier (e.g. drlig carrier system) or inert carrier, slich as a glycoside.

Brief Description of the Drawings:
Figlire 1: Headspace analysis lising GC-MS of wild (A) and cliltivated strawberry (B). Chromatograms are of ni/z 93 (obtained in SCAN mode). Peaks; 1, α-pinene; 2, β-myrcene; 3, β-phellandrene; 4, γ-terpinene (tentative); 5, α-terpinolene (tentative); 6, α-terpineol; 7. carvylacetate; 8, linalool; 9, trans-nerolidol.
Figlire 2: Seqlience alignment of H64 proteins H64NORS, H64MliT, H64VES. H64MliT nlicleic acid seqlience after the stop codon in H64NORL (location marked) was changed into a Lelicine residlie. Black backgrolind corresponds to identical residlies in all three seqliences and gray backgrolind corresponds to identity between two olit of the three seqliences.
Figlire 3: targeting signals in the different H64 genes.
A. Protein seqlience alignment of the different H64 fragments obtained by PCR on
genomic DNA and the same regions in cDNAs isolated. Arrows indicate which one
of the seqliences do not have a stop codon in this region. The RR motif is common
in targeting signals of monoterpene synthases. Black backgrolind corresponds to
identical residlies in all seven seqliences and gray backgrolind corresponds to
identity between at least three olit of the seven seqliences.
B. Site directed mlitagenesis and constrlicting H64MliT from H64N0RL. The
5'region of H64NORL and H64MliT is aligned between the two ATG codons and
the position of directed mlitagenesis is marked by the gray backgrolind. The stop
codon T(li)GA in H64N0RL was converted to a codon encoding a lelicine residlie
(CTA).
Figure 4: Expression of H64 genes analyzed by RNA gel blots and H64NORL cDNA as a probe.
A. Expression in vegetative (leaves) and reprodlictive (4 stages of frliit development) tisslies.

B. Expression in ripe frliit of two wild cliltivars (1, Plant Research International line
HI and 2 Plant Research International line 92189) and two cliltivated cliltivars (1.
cliltivar Calypso and 2, cliltivar Gorrella.
C, Expression in frliits treated with or witholit the synthetic alixin NAA. Strawberry
frliit (cliltivar Elsanta) at the white stage of development were treated with
lanolin paste containing lOOmM NAA. Treated and control berries (paste with no
NAA) were treated, left on the vine for 7 days and then picked and lised for RNA
isolation.
Figlire 5: The pRSET B expression vector lised for cloning and expression of H64MliTA SOSA and SOSV in E.Coli cells.
Figlire 6: Nlicleic acid seqlience alignment of the two cliltivated SOSAcDNAs cloned (MA and WS) and their homolog from the wild strawberry (SOSV). Black backgrolind corresponds to identical residlies in all three seqliences and gray backgrolind corresponds to identity between two olit of the three seqliences. The location of the CC insertion calising the frame shift and the stop codon following it is depicted. The stop codon at the 3' is the end of the ORF.
Figlire 7: Alignment of the protein seqlience of the different SOS cDNAs isolated. Black backgrolind corresponds to identical residlies in all five seqliences and gray backgrolind corresponds to identity between at least three olit of the five seqliences. The insertion of CC in SEQ11B(S0SA/MA) and SEQ10B(SOSA/WS) forms a proline residlie and a stop codon after. In SEQllC(SOSA/MA) and SEQ10C(SOSAAVS) the two cytosine nlicleotides were removed and allowed flirther translation of the protein.
Figlire 8: Nlicleic add seqlience alignment of the different SOS fragments obtained by PCR on genomic DNA and the same regions in cDNAs isolated from wild and cliltivated strawberry. The solirce of the fragment is marked in the left side of each seqlience name. Black backgrolind corresponds to identical residlies in all fifteen seqliences and gray backgrolind corresponds to identity between at least twelve olit of the fifteen seqliences.

Figlire 9: Expression of SOS genes analyzed by RNA gel blots and SOSV cDNA as a
probe
Expression in ripe frliit of two wild cliltivars (1, Plant Research International lins. H1
and 2 Plant Research International line 92189) and two cliltivated cliltivars (1,
cliltivar Calypso and 2, cidtivar Gorrella.
Figlire 10: Radio-GLC analysis of radio-labelled prodlicts formed from [3H]-geranyl diphosphate in assays with recombinant proteins. A, FID signal showing linlabelled alithentic standards of 1, 6-myrcene; 2, trans-ocimene; 3, linalool; 4, α-terpineol; 5, nerol; 6, geraniol. B,C, radio-traces showing enzymatic prodlicts of recombinant proteins SOSV (B) and H64MliT (C).
Figlire 11: Radio-GLC analysis of radio-labelled prodlicts-formed from [3H]-farnesyl diphosphate in assays with recombinant protein. A. FID signal showing linlabelled alithentic standards of 7, cis-nerolidol; 8, frans-nerolidol; 9, trans-trans-farnesol. B, radio-trace showing enzymatic prodlicts of recombinant protein H64JVIliT.
Figlire 12: GC-MS analysis on an HP5-MS collimn of prodlicts formed from geranyl diphosphate in assays with recombinant SOSV protein. Peaks: 1, α-pinene; 2, β-pinene; 3, sabinene; 4, β-myrcene; 5, α-phellandrene; 6, D-phellandrene; 7, dihydromyrcenol (tentative); 8, α-terpinolene (tentative); 9, α-terpineol (tentative),
Figilire 13: GC-MS analysis on an HP5-MS collimn of the prodlict formed from geranyl diphosphate in an assay with recombinant H64MliT protein. A, wz/r 93 chromatogram. B, mass spectrlim of the major prodlict peak (linalool).
Figlire 14: GC-MS analysis on an HP5-MS collimn of the prodlict formed from farnesyl diphosphate in an assay with recombinant H64MliT protein. A, m/z 93 chromatogram. B, mass spectrlim of the major prodlict peak (nerolidol).

Figlire 15: Transient GFP expression of flision proteins in tobacco protoplasts, g, GFP; ca, clilorophyll alito-flliorescence; mt, MitoTracker (mitochondrial stain); ol, overlay of chlorophyll alito-flliorescence image and GFP image; ol-mt, overlay of chlorophyll alito-flliorescence image, GFP image and Mitotrackcr image. 10 diffeirent constrlicts were made (C1-C10) to stlidy fragments derived from H64NORL (Cl, C2), H64TAR4 (C3. C4. C5) and H64VES (CT, C8, C9). See Figlire 16 for a schematic representation of the different constrlicts made and lised for the localization stlidies. C6 shows localization effusion of a citrlis limonene synthase 5' end with GFP. C10 is a flision of the H64VES region between the two Methionine residlies and the region down stream of the second Methionine from H64NORL. pOL65 is the original vector, containing only GFP and was lised to insert all fragments for flision with the GFP. Rpo-ol is a positive control for plastidic targeting signal. Chloroplasts are on average 5 micrometer in size while mitochondria are 1 micrometer in size. pOL65, Cl, C2, C4, C5, C8 and C9 all show cytosolic localization. C3 shows dlial plastidic and mitochondrial localization. C6, C7. C10 and Rpo-ol show plastidic sliβ-celllilar locahzation.
Figlire 16: Schematic representation of the different constrlicts lised for GFP transient expression assays in tobacco protoplasts. Fagments derived from the 5'-end of the cDNAs described in the invention were lised for a translational flision with the GFP gene. The MID motif is present in most sesqliiterpene synthase genes described lip to date. SC, stop codon. Ml and M2 are the two methionine residlies at the N-termini of the variolis proteins (see also Figlire 3A).
Figlire 17: Comparison of effects of famesol and hnalool present in the growth medilim on mycelilim growth of Phytophthora infestans.
Figlire 18; Dose-response data of effects of linalool present in the growth medilim or the vapolir phase on mycelilim growth of Phytophthora infestans.
Figlire 19: Dose-response data of effects of nerolidol present in the growth medilim or the vapolir phase on mycelilim growth of Phytophthora infestans.

Figlire 20: Dose-response data of effects of linalool and nerolidol present in the growth medilim alone and in combination on mycelilim growth of Phytophthora infestans.
Figlire 21: Dose-response data of effects of linalool and nerolidol present in the growth medilim alone and in combination on mycelilim growth of Phytophthora infestans.
Figlire 22
Dose-response data of effects of linalool and nerolidol present in the growth medilim
on mycelilim growth of Flisarilini spp. on day 7.
Figlire 23
Dose-response data of effects of linalool and nerolidol present in the growth medilim
on mycelilim growth of Botrytis spp. on day 7.
Figlire 24
Dose-response data of effects of nerolidol (A) and linalool (B) present in the growth
medilim on spore germination of Flisarilim verticillioides isolates on day 3.
Figlire 25
Headspace analysis of transgenic Arabidopsis expressing the H64NORS with the H64VES targeting signal (H64TAR) cDNA, showing a large peak of linalool (1). and a smaller peak of nerolidol (2). Both compolinds are absent in control, wildtype Arabidopsis (see insert).
Figlire 26
Headspace analysis of volatiles prodliced by control and transgenic, H64TAE expressing, potato (3 individlial transformants TM 9, TM 13, TM 29). Linalool is virtlially absent in control potato, and strongly enhanced in the transgenic lines. Also 8-hydroxylinalool is enhanced in the transgenic lines.

Figlire 27
Chii-al analysis of the free linalool in control and transgenic potato, showing the presence of both enantioraers in control potato (abolit 80:20). In the H64TAR trangenic lines the ratio has shifted drammatically to the S-enantiomer, that is prodliced by the introdliced enzyme.
Figlire 28A
Identification of linalyl-βi-D-gllicopyranoside in Petlinia tisslie lising HPLC-MS/MS. Ion trace m/z 375 of A: the synthesized (i?,S)-linaly-β-D-gllicopyranoside, B: the transgenic Petlinia leaf tisslie and C: The control Petlinia leaf tisslie.
Figlire 28B
Prodlict ion spectrlim of A: The synthesized (/?,S)-linalyI-fi-D-gllicopyranoside and B: The compolind isolated from the transgenic Petlinia tisslie. Retention time and prodlict ion spectrlim of the synthesized (/2,S)-linalyl-β-D-gllicopyranoside fit with the compolind detected in the transgenic Petlinia tisslie.
Figlire 29
Determination of the enantiomeric distriblition of S- and i2-linalool after enzymatic hydrolysis of the gllicoside firaction obtained fi-om leaf tisslie lising chiral phase MDGC-MS analysis, A: Control tisslie and B: the transgenic tisslie. The transgenic plant acclimlilates higlily enriched S-linalyl-β-D^gllicopjrranoside.
Figlire 30
Figlire 30 combines the data of table 1 and 4. Figlires 30 A, B and C provide the correlation in lesion size, lesion growth rate, and sporidation respectively of Phytophthora infestans isolate IPO 428-2 plotted against the content of linalool, 8-hydroxylinalool, linalooltriol, lynalylgllicoside, 8-hydroxylinalylgllicoside and linalyltriolgllicoside content of the potato transgenic lines T or TM-9, -13, -29 and a

control line. The control data from table 4 on flingal growth and sporlilation were taken to be the average vallies of the H64NOR plants with negligible increased level of either linalool, nerolidol or derivatives. The linalool (derivative) data provided in table 1 are mlich more reliable and qliantitative than the SPME data on linalool in table 4, which jlistifies their lise. Figlire 30 D provides the in vitro data on the sensitivity of Phytophthora infestans isolate IP0428-2 which was lised for the in planta experiments to plire linalool in the medilim as described in Example 9.

Examples
The following examples are offered by way of illlistration.
Example 1. Analysis of terpenes in wild and cliltivated strawberry
Terpenoid Biosynthesis in Wild and Cliltivated Strawberries
The cliltivated variety (Elsanta) lised by lis for the mentioned experiments prodlices both the monoterpene linalool and the sesqliiterpene nerohdol. On the other hand the wild cliltivar lised (PRI line 92189) prodlices low levels of linalool blit does not show a trace of nerolidol. Both literatlire reports and olir own GC-MS data show similar patterns of linalool and nerolidol prodliction in several other cliltivated and wild strawberry varieties. Olir seqliencing data and experiments lising the recombinant enzymes prodliced in Exoli show that the capability of the cliltivated variety to form nerolidol was acqliired by removing (by deletions and translation stop) the targeting signal to the plastid [were the slibstrate for monoterpene biosynthesis is available (GPP)] and by directing the translation start to the downstream AliG codon. However, linalool in the cliltivated varieties may also be formed by enzymes encoded by genes similar to H64TAR2, H64TAR4 and H64TAR6 which contain a proper targeting signal with no stop and therefore their protein prodlicts are directed to the plastid for forming linalool. If GPP is present in the cytosol, then linalool colild also be prodliced there by an enzyme encoded by a cytosolically expressed cDNA. We can not excllide that translation in H64TAR2, H64TAR4 and H64TAR6 may also start from the downstream AliG codon (the one downstream from the RR motif and not the additional AliG codon present jlist prior the RR motif) and this will reslilt in the formation of nerolidol as well. However, since cliltivated varieties like the ones lised in this stlidy are mostly octaploids it is likely that evollitionary processes as polyploidity allows the plant to form an additional (mlitated) gene from an existing gene and to prodlice an additional beneficial compolind slich as nerohdol for flavolir and defense. Williams et aL, (Biochemistry 1998,37,12213-12220) described a role for the tandem arginines present in the N-terminal of monoterpene synthases in the

liniqlie diphosphate migration step accompanying formation of the intermediate 3-s-linalyl diphosphate and preceding the final cyclization reaction catalyses by the monoterpene synthases. This RR motif is present in H64TAR2, H64TAR6, and H64VES and this might explain the formation of linalool by this genes encoding en2ymes. However, the H64MliT recombinant protein does not contain the RR motif blit catalyses the formation of both nerolidol and linalool. This might implicate other residlies between the RR motif location and the down stream AliG as flinctioning to determine whether monoterpene will be formed.. This motif contain 12 amino acids: N-termini- DSLLPSSITIKP.
The short genomic DNA seqlience obtained (H64W149) contains an RW motif instead of an RR motif and it might be of importance for the formation of the monoterpene linalool. In the wild cliltivars (diploid) only one variant encoding a protein with a targeting signal colild be identified (both by PCR on either DNA and RNA) which may only catalyze the formation of the low levels of linalool detected.
Headspace analysis. Samples of ripe or ripening frliits were enclosed in 1-L glass jars that were closed with a teflon-lined lid eqliipped with an in- and olitlet, and placed in a climate room at 25oc and 210 μmol.m-2 provided by 400-W HPI-T lights (Philips, Eindhoven, the Netherlands). A vaclilim plimp was lised to draw of air throligh the glass jars at approximately 100 mL min-1 with the incoming air being plirified throligh a glass cartridge (140 x 4 mm) containing 150 mg Tenax TA (20/35 mesh, Alitech, Breda, the Netherlands). At the olitlet the volatiles emitted by the firliits were trapped on a similar Tenax cartridge. Volatiles were sampled dliring 24 h. Cartridges were ellited lising 3x1 mL of redistilled pentane-diethyl ether (4:1). Of the (non-concentrated) samples, 2μL were analysed by GC-MS lising an HP 5890 series II gas chromatograph eqliipped with an HP-5MS collimn (30 m x 0.25 nam i.d., 0.25 μm df) and an HP 5972AMass Selective Detector. The GC was programmed at an initial temperatlire of 45°C for 1 min, with a ramp of 10o min-1 to 280°C and final time of 5 min. The injection port (sphtless mode), interface and MS solirce temperatlires were 250, 290 and 180oC, respectively, and the He inlet presslire was controlled by electronic presslire control to achieve a constant collimn flow of 1.0 mL

min-1 Ionization potential was set at 70 eV, and scanning was performed from 48-250 amli.
The analysis of the headspace profiles was foclised on terpenoids by only showing the ion 93 chromatogram (altholigh samples were analysed lising the SCAN mode). In that way, remarkable differences can be seen between cliltivated and wild strawberry: the headspace profile of the wild strawberry contains carvylacetate and a nlimber of olefinic monoterpenes slich as α-pinene, myrcene, α-phellandrene, and α-terpinolene, α-terpineol and γ-terpinene (the last three tentatively identified) (Figlire lA), whereas the cliltivated is dominated by two major peaks only: linalool and trans-nerolidol (Figlire IB).
Example 2. General Moleclilar Techniqlies
DNA was isolated fr^m yoling strawberry leaves as described by Marty et al., [Theor. Appl. Genet (2000) 100:1129-1136].
RNA gel blots experiments were performed as described by iVharoni et al., [The Plant Celi, (2000) 12, 647-661].
Cloning flill length cDNAs was performed by lising the SMART RACE cDNA Amplification Kit (Clontech) according to the manlifactlirer instrlictions with slight modifications either to annealing temperattires (normally redliced by 5 to 10oC compared to the one recommended) or amolint of cycles (lip to 35 cycles). PCR, restriction digests, plasmid DNA isolation and gel electrophoresis were performed lising standard protocols. All fragments were plirified olit of gel lising the GFX plirification kit (Amersham). (Zoning of PCR fr-agments was either done to the PCR SCRIPT (Stratagene) or pCR 4Bllint-T0P0 (Invitrogen) vectors (for bllint end prodlicts generated when lising pfli polymerase) or to the pGEM-T Easy (Promega) vector (when A tailed PCR prodlicts were generated by the lise of taq polymerase). Throligholit the text the following constrlict/cDNA names will be lised (also see seqlience listing):
- H64VES: wild strawberry, flill length cDNA (with targeting signal)
- H64N0RL: original cliltivated strawberry cDNA starting from Met 1, inclliding stopcodon between Met 1 and Met 2 (non-flinctional targeting signal)

- H64NORS: derived from H64NORL starting from Met 2 (no targeting signal)
- H64MliT: derived from H64NORL; stopcodon repaired
- H64TAR: lised for transformation of plants: composed of H64VES Metl to Met 2 region and H64NORS (with targeting signal)
- H64NOR: lised for transformation of plants: H64NORS inclliding intron
Example 3. Constrliction of a Strawberry Red Frliit Stage cDNA Library, Mass Excision and Random Seqliencing
Messenger RNA Isolation and cDNA Library Constrliction
Total RNA was isolated from strawberry frliit red stage of development lising the method described by Manning K. [Analytical Biochemistry (1991) 195, 45-50]. The cliltivar lised was Fragaria X ananassa Ditch, cv. Elsanta. The cDNA library was prodliced as a clistom service by (Stratagene) in the lambda zap vector. Messenger RNA was isolated from total RNA lising the polyA+ isolation kit (Pharmacia).
Mass Excision and Random Seqliencing
The ExAssistTM/SOLRTM system (Stratagene) was lised for mass excision of the pBlliescript SK(-) phagemid. The excision was done according to the manlifactlirer instrlictions lising 20μ1--3pfli from the non-amplified library for each excision. High qliality plasmid DNA from randomly picked colonies was extracted lising the QIAGEN BioROBOT 9600. Colonies were grown overnight in 3ml Lliria Broth medilim (10g/l tryptone, 5 gA yeast extract, 5 g/1 NaCl) slipplemented with 100 mgA ampicillin, centrifliged at 3000 RPM for 10 min. and the pellet was lised directlf for plasmid DNA isolation by the robot. Each DNA isolation rolind consisted of 96 cliltlires.
Insert size was estimated by agarose gel electrophoresis after restriction enzyme digestion of the pBllieScript (SK-) vector with EcoRI and XhoL Inserts with length above 500 bp were lised for seqliencing. Plasmid DNA from the selected samples were lised for polymerase chain reaction (PCR) seqliencing reactions lising the ABI PRISM™ Dye Terminator Cycle Seqliencing Ready Reaction Kit and the MJ Research PTC-200 DNA Engine™ thermal cycler. The T3 and T7 liniversal primers

were lised for seqliencing from the 5' and 3'ends respectively. PCR program was according to the Dye Terminator manlifactlire's protocol (ABI PRISM). The ABI 373, 370A and 310 seqliencers (Applied Bio-systems) were lised. Seqliences were edited manlially to remove vector and non reliable seqliences and slibmitted to the BLAST homology search (Altschlil et al. J. Mol. Biol. 215, 403 - 410, 1990) provided by the National Center for Biotechnological Information on the world wide web (infQ@ncbi.nlm.nih.gov). Search was performed against ali non-redlindant data bases available by the program.
Example 4. Cloning and Characterization of H64 Genes from Wild and Cliltivated Strawberry
Cloning of the H64 cDNA from Cliltivated Strawberry (H64NORL) and its Homologlie from the Wild Strawberry (H64VES)
We primarily identified the H64 cDNA olit of olir randomly seqlienced clones originating from the cliltivated strawberry cliltivar Elsant-a (ripe red frliit) cDNA library. Homology search reslilts lising the BLAST program indicated that the cDNA might encode a terpene synthase protein. The entire H64 cDNA is 1874 bp long [(termed H64 Normal Long (H64NORL)] and contains a open reading frame (ORF) encoding a 519 amino acids (aa) long protein [we termed the part of the cDNA forming the 519 aa ORF as H64 Normal Short (H64NORS), see Figlire 2].
Cloning of the wild strawberry homolog of the cliltivated H64 cDNA was accomplished by the lise of the SMART RACE kit (Clontech) lising RNA from the Plant Research International collection of wild strawberries (line 92189). Oligonlicleotides primarily lised for seqliencing the H64NORL cDNA were lised for 3' RACE amplification (AAP291 - 5'- CTTCATGAGGTTGCACTTCG- 3' and the nested oligonlicleotide AAP 293 - 5'- AATGGTGGAAGGAGCTTGKIATTGG- 3'). The flill length wild strawberry cDNA [H64 Vesca (H64VES)] was obtained by designing an oligonlicleotide on the 3' lintranslated region (liTR) based on the 1000 bp fragment obtained in the 3' RACE and lising it to RACE for the 5' side (5' GTTCAACTCCACnTCCAGCAGTC 3'). The H64VES cDNA is 1894 bp long and contains a open reading frame encoding a 580 anlino acids (aa) long protein. Sixty

one amino acids downstream of the first methionine residlie of the 580 aa protein we colild identify an additional methionine residlie. This 61 amino acids resemble the characteristic plastidic targeting signal of monoterpene synthases since it contains the two arginines motif and a large nlimber of serine residlies [Williams et al. (Biochemistry, 37 12213-12220, 1998); see Figlire 3A). The H64NORL and H64VES cDNAs share 96 % identity at the nlicleic acid level and if the stop codon is eliminated and the rest of the seqlience translated, 92.4 % at the amino acid level (from the ATG located at nlicleotide 145 lip to the end of the coding region). H64VES and H64NORS share 97.2 % identity at the nlicleic acid level and 94.2 % at the amino acid level (when the part starting from the beginning of H64NORS from H64NORVES is lised for the ahgnment lip to the end of the coding region).
Analysis of H64 Expression Dliring Development, in Cliltivated and Wild Cliltivars and in Response to Alixin Treatment
RNA gel blot analysis lising H64NORL as a probe revealed that it is lipreglilated dliring the cliltivated strawberry frliit ripening (Figlire 4). No expression colild be detected in the leaf and green frliit tisslies. H64 expression increased from the white to red stage of frxdt development. Analysis of H64 expression in ripe frmts of two wild and two cliltivated cliltivars showed that H64 is strongly expressed in the cliltivated cliltivars and hardly any expression colild be detected in the wild cliltivars (slight signal was detected in the wild cliltivars after long exposlire of the film, data not shown). Another RNA gel blot showed that H64 is repressed by alixin. This correlates with the fact that also other ripening lip-reglilated genes in strawberry are repressed by alixin.
Site Directed Mlitagenesis of H64NORL
A more thoroligh analysis of the H64 cDNA (termed H64NORL) revealed that it might contain an additional ATG start codon, 99 bp lipstream of the original ATG we identified (proposed to be the beginning of the ORF encoding the 519 aa H64NORS protein). The two ATG codons were located in frame blit no peptide colild be formed between them since a stop codon located 39 bp before the down stream ATG was evident. We slispected that the part between the two ATG is actlially part of the

protein and for some reason it might be mlitated so a shorter protein starting for the downstream ATG might be formed. Additional slipport to this idea was the high ablindance of serine residlies identified in the translated area between the two ATGs. It resembled N-termini of other monoterpenoid synthases which contain relativelγ-high ablindance of serine residlies. We therefore employed site directed mlitagenesis in order to modify the stop codon and constrlict a non trlincated H64N0RL protein [termed H64 Mlitagenized (H64MliT)1. By changing the stop codon (TGA) into a lelicine residlie (CTA) the H64MliT cDNA is 1659 bp long containing a 552 aa long protein (see Figlire 3B). The site directed mlitagenesis was performed lising the QuikChange kit as described by the manlifactlirer (Stratagene). The ohgonlicleotide lised for the exchange was. 5' GGGAAGCAAGCTATCTAGAAAGTAGCAG-GCAATT-3'.
PCR on Cliltivated Strawberry Genotnic DNA
In order to verify whether the seqlience we obtained for H64NORL was not a PCR artifact and the stop codon between the two ATGs exists, we performed PCR on the cliltivated strawberry genomic DNA. We designed two ohgonlicleotides one lipstream the first ATG (5'- CTCCCACAGCTTCTTAGTTGC- 3') and the other downstream of the second ATG (the beginning of H64NORS) (5-
CTAGCTCTGCTTACATTCCrrCAAGAC- 3'). Amplification with these two oligonlicleotides was expected to amplify a fragment of approximately 200 bp. We obtained two clear firagments of 300 bp and 400 bp each. Seqliencing folir clones of the 300 bp length firagments revealed them to be similar to the original H€4N0RL cDNA. Seqliencing and aligning 20 of the larger clones identified several isoforms which were different firom the original cliltivated H64NORL cDNA. Ali fi-agments (inclliding the short ones) contained an intron of approximately 100 bp. Fom' liniqlie different clones olit of the 20 seqlienced were identified. Two of them [SEQ6C(H64N0Rli1/W151) and SEQ7C(H64NORV2nJP3)] had an additional 20 aa (compared to H64MliT) blit still contained a stop codon located immediately at the beginning of the peptide they formed. Other two firagments
[SEQ8C(H64N0Rli3/liP16) and SEQ9C(H64N0Rli4/liP1)] did not contain any stop codon and were most similar to the seqlience of H64VES. These fragments added 26

aa to the H64MliT seqlience and they both contain the two arginine residlies as in H64VES which are most often folind in the plastidic targeting signal of monoterpene synthases (see Figlire 3A).
Cloning H64MliT/H64NORS for Expression in E,coli
The Exoli expression vector pRSETB (Invitrogen) was lised for heterologolis expression of strawberry terpene synthases (see Figlire 5). The pRSETB vector contains the T7 promoter which can be indliced by isopropyl-p-D-thiogalactopyranoside (IPTG) and therefore by inserting the desired gene downstream of this promoter, the gene can be expressed in E. coll. In addition, DNA inserts were positioned downstream and in fi:ame with a seqlience that encodes an N-terminal flision peptide. This seqlience incllides (in 5' to 3' order from the N-terminal to C-terminal), an ATG translation initiation codon, a series of six histidine residlies that flinction as a metal binding domain in the translated protein, the Anti-Xpress epitope, and the enterokinase cleavage recognition seqlience.
CP
t>
The original pRSETB was primarily lised for the insertion of the gene encoding the Green Flliorescent Protein (GFP). The GFP gene was flised to the pRSETB vector lising the BamHI and Hindlll restriction sites located at the mliltiple cloning site (MCS) as can be seen in figlire 5. This constrlict for the expression of GFP served as control for the experiments together with the empty pRSETB vector. Cloning the GFP gene to the pRSETB vector inserted an additional Sail restriction site at the 3' of the GFP gene and together with the BamHI site located at the 5' of the GFP gene served as sites for cloning H64MliT- The BamHI and Sail sites were introdliced to the 5' and 3' respectively of the H64MIJT coding seqlience by the lise of PCR, The 552 amino acid open reading firame of the H64MliT clone was amplified with the pfli DNA polymerase (Stratagene) and oligonlicleotides (containin the BamHI and SaII sites) AAP339 (5'- CGGATCCGGCATC-GTCTTCTCGGGC- 3') and AAP334 (5'- CGTCGACCAACTCCACTTCCGGTAGTC- 3') according to the manlifactlirers instrlictions. The PCR prodlict was cloned into PCR-script vector (Stxatagene), clit olit with BamHI and Sall and flirther inserted (as a translation flision) into the corresponding restriction sites in the pRSETB vector. H64NORS was cloned in a similar way.

Bacterial Expression and Partial Plirification lising the His tag Collimns. The pRSETB vector harboring the H64MliT or H64NORS was lised to transform E. coli strain BL21 Gold DE3 pLysE (Stratagene) as described by the manlifactlirer. For bacterial expression typically 1 ml of overnight liqliid cliltlire gi'own at 37°C in Lliria Broth (LB) medilim (lOgA tryptone, 5 gfl yeast extract, 5 gA NaCl) slipplemented with 100 mgfl ampicillin was dillited 50 times in the same medilim and grown lintil the OD600 reached 0.4 (at 37o C). At this stage IPTG was added to a final concentration of ImM in order to indlice expression . After overnight growth at 16°C the cells were harvested by centrifligation at 4000 x g for 15 min. Pellet and a sample from the slipernatant were kept for SDS gel analysis. The cells were flirther processed as described by the Ni-NTA Spin Collimns manlifactlirers (QIAGEN) for protein plirification linder native conditions. First ellite from the collimn (200 fil) was flirther lised for enzymatic activity assays.
Example 5. Cloning and Characterization of SOS Genes from Wild and Cliltivated Strawberry
Cloning of the SOS cDNA from Cliltivated Strawberry (SOSA) and its hom,olog from the Wlid Strawberry (SOSV)
For cloning the SOSA(MA) cDNA from the cliltivated strawberry CV Elsanta, we designed an oligonlicleotide on a pliblished seqlience of a sesqliiterpene cyclase fi-om the wild strawberry (Nam et al. Plant MoL Biol. 39: 629-636, 1999). The oligonlicleotide (AAP 272, 5'- GATGATATGTATGATGCATTCGG- SO was lised to perform a 3' RACE reaction lising the RACE kit (Clontech) and a 991 bp fi-agment was cloned. For cloning the flill length cDNA we performed a 5' RACE reaction lising an oligonlicleotide designed on the 3' liTR of the cDNA (AAP283, 5'-GAAAGGATAGGCTCATCAGTACGTG- 3'). The entire S0SA(1^IA) cDNA cloned is 2605 bp long. We however colild not identify an ORF encoding a protein longer then 255 aa, which is less then a half of a tjqpical terpene synthase. Therefor a second attempt to clone a cDNA with a longer ORF was performed. lising oligonlicleotides based on the SOSA(MA) seqlience, one located on the beginning of the ORF (AAP325,

5'- CGGATCCGCCTGTCCATGCTACTCC" 3') and the other on the liTR (AAP341, 5'-CGTCGACTGAGTTCAGAGTGGCACTGG- 3'), a second full-length SOSA cDNA was isolated by the means of PCR on the cliltivated strawberry cDNA [termed SOSA(WS)]. Seqliencing SOSA(WS) revealed that as for SOSA(MA) it contains a trlincated ORF. We decided to clone the flill length SOS homolog from the wild strawberry in order to identify the calise for slich a trlincation in the cliltivated genes ORF. Cloning of the wild SOS homolog was performed by 3' RACE reaction lising an ohgonlicleotide designed on the SOSA(MA) ORF (AAP325, see above). The flili length SOS homolog from the wild strawberry (SOSV) is 1973 bp long and contains a ORF encoding a 556 aa long protein. Aligning SOSA(MA), SOSA(WS) and SOSV nlicleic acid seqliences revealed minor changes in the ORF (see Figlire 6). We colild however identify the basis of the trlincation in the cliltivated SOS genes which was an insertion of two cytosine nlicleotides calising a frame shift followed by a stop codon (see Figlire 6). Removing the CC insertion from the SOSA(WS) and SOSA(MA) genes reslilts in the formation of ORFs encoding 554 and 555 aa respectively (Figlire 7).
PCR on Cliltivated and Wild Strawberry Genomic DNA
In order to confirm the presence of the CC frame shift, calising a trlincation in the cliltivated strawberry SOS genes we analyzed the existence of the insertion at the DNA level PCR on both wild and cliltivated strawberry genomic DNA was performed lising two oligonlicleotides located from both sides of the place of insertion (AAP345, 5'- AGAGGTTAGKJTGCrrCCJGCGTTAC- Z') and the reverse ohgonlicleotide, AAP346, 5' GAACAACTCCACGATCCTrATCrrC- 3^. The expected amplified DNA Segment was 200 bp. PCR prodlicts at the size of 300 bp were obtained from both reactions lising the wild and cliltivated DNA We seqlienced 20 and 15 fragments from the cliltivated and wild strawberry reactions respectively. AH fragments contained an intron of approximately 100 bp. Seqlience alignment of all fragments revealed 7 different seqliences from the cliltivated and 5 from the wild. Figlire 8 shows an alignment of all fragments of the SOS genes both from the wild and cliltivated strawberry obtained either from RNA (the different cDNAs) or from DNA. Among the cxiltivated fragments we colild identify 2 fragments which showed the CC insertion while the other 5 did

not contain it. On the other hand no fragment in the wild strawberry colild be detected that contained the frame shift mlitation.
Analysis of SOS Expression in Ripe Cliltivated and Wild Strawberry Frliit lising the SOSV cDNA as a probe we analyzed SOS gene expression in two different wild and cliltivated cliltivars (Figlire 9). The SOSV cDNA colild be lised for hybridization with blots containing RNA from both wild and cliltivated cliltivars since the SOSA genes and SOSV share nearly 99 % identity at the nlicleic acid level (in the ORF region). Hardly any expression colild be detected in the cliltivated cliltivars while strong expression colild be detected in the wild cliltivars. The SOSA probe was also lised for hybridizing blots with RNA extracted from different cliltivated (Elsanta) frliit developmental stages, blit jlist weak signal colild be detected after long exposlire. Nam et al., (1999) were also not able to detect expression of the partial cDNA homolog of SOS with RNA derived from different frliit developmental stages of the cliltivated strawberry. Expression in different wild strawberry plant tisslies was restricted to the frliit, specifically to the red ripe stage.
Cloning and Expression of SOSV and SOSA in E.coli
Both the SOSA and SOSV coding regions were lised for the formation of a recombinant protein in E,Coli cells. The entire ORF of SOSA cDNA altholigh trlincated was expressed in order to serve as a negative control for the enz3rmatic assays. Similar to the cloning of H64MliT the BamHIand Sail restriction sites at the 5' and 3'of the GFP gene respectively served as sites for the cloning of SOSA and SOSV ORFs into the pRSETB expression vector. The BamHI and SaZI sites were introdliced to the 5' and 3* respectively of the wild and cliltivated SOS genes coding seqlience by the lise of PCR. The restriction sites were added to the oligonlicleotides lised for PCR reaction (AAP325, 5^ CGGATCCGCCTGTCCATGCTACTCC- 3' and the reverse primer AAP341, 5'- CGTCGACTGAGTTCAGAGTGGCACTGG- 3'). The PCR prodlict was cloned into PCR-script vector (Stratagene), clit olit with BamHIand Sail and flirther inserted (as a translation flision) into the corresponding restriction sites in the pRSETB vector. Expression of SOSA and SOSV in E. Call was performed parallel to the expression of H64MliT and linder identical experimental conditions.

Example 6. Analysis of SOSA, SOSV, H64MliT and H64NORS recombinant enzymes
For determination of terpene synthase identity, the His-tag plirified enzymes (prepared as described above linder Example 4.6) were dillited 10-fold with bliffer A containing 15 mM MOPSO (pH 7.0), 10% glycerol, 10 mM MgCb, 1 mM sodilim ascorbate and 2 mM DTT. To 1 mL of this enzyme preparation, 40μM of either PH]-geranyl diphosphate (GPP) or pH]-farnesyl diphosphate (FPP) were added. Assays with GPP as slibstrate were also slipplemented with 1 mM MnCl2. After the addition of a 1-mL redistilled pentane overlay, the tlibes were careflilly mixed and inclibated for 1 h at 30oC. After the assay, the tlibes were vortexed, the pentane layer was removed and passed over a short collimn of alliminlim oxide overlaid with anhydrolis Na2SO4. The assay was re-extracted with I mL of diethyl ether, which was also passed over the alliminlim oxide collimn, and the collimn washed with 1.5 mL of diethyl ether. 100 jiL of the organic exrtract was removed for liqliid-scintillation colinting in 4.5 mL of scintillation cocktail (liltima Gold. Packard Bioscience. The Netherlands). Radio-labelled prodlicts were present in the organic extracts of:
H64MliT H64NORS SOSV SOSA
____ ^ ^ ^
PHl-FPP + + - ,
Slibseqliently, the extracts were carefidly concentrated linder a stream of No before analysis lising radio-GLC and GC-MS. Radio-GLC was performed on a Carlo-Erba 4160 Series gas chromatograph eqliipped with a RAGα-90 radioactivity detector (Raytest, Stralibenhardt, (Germany). Sample components elliting from the collimn were qliantitatively redliced before radioactivity measlirement by passage throligh a conversion reactor filled with platinlim chips at 800°C. Samples of 1μL were injected in the cold on-collimn mode. The collimn was a flised silica capillary (30 m x 0.32 mm i'd.) coated with a film of 0.25 fim of polyethylene glycol (EconoCap EC-WAX, Alltech Associates) and operated with a He-flow of 1.2 mL min-1. The oven temperatlire was

programmed to 70oC for 1 min, followed by a ramp of 5° min-1 to 210oC and a final time of 10 min. Abolit 20% of the collimn effllient was split with an adjlistable splitter to an FID (temperatlire 270oC). The remainder was directed to the conversion reactor and radio detector. H2 was added prior to the reactor at 3 mL min-1, and CH4 as a qliench gas prior to the radioactivity detector (5 mL colinting tlibe) to give a total flow of 36 mL min-1, Radio-GLC analysis gave the following reslilts: - the SOSV and H64MliT and H64NORS recombinant proteins catalysed the
formation of radio-labelled prodlicts from PH]-GPP (Fig 10). For the SOSV protein a nlimber of radio-labelled prodlict peaks were visible in the retention time area of olefinic monoterpenes (Fig. lOB). The major radio-labelled prodlict did not co-ellite with any of the added linlabelled reference compolinds, blit one of the minor radio-labelled peaks seemed to co-ellite with the reference β-myrcene. For the H64MliT recombinant enzyme the single radio-labelled prodlict co-ellited with linalool (Fig. 100).
with PH]-FPP as slibstrate scintillation colinting showed that neither the SOS A nor the SOSV recombinant protein catalysed any radio-labelled prodlict formation. The H64MliT protein catalysed the formation of a radio-labelled prodlict which radio-GC analysis showed to be one single prodlict, co-elliting with traas-nerolidol (Fig. 11). The samples were also analysed by GC-MS lising a HP 5890 series II gas chromatograph eqliipped with an HP5-MS collimn (30 m x 0.25 mm Ld., 0.25 μm di) and HP 5972A Mass Selective Detector (Hewlett-Packard). The oven was programmed at an initial temperatlire of 45°C for 1 min, with a ramp of 10°C min-1 to 280°C and final time of 5 min. The injection port (splitless mode), interface and MS solirce temperatlires were 250, 290 and 180°C, respectively, and the He inlet presslire was controlled by electronic presslire control to achieve a constant collimn flow of 1.0 mL min-1 Ionization potential was set at 70 eV, and scanning was performed from 48-250 amli. The m/z 93 chromatogram of SOSV recombinant protein catalysed prodlicts from PH]-GPP again shows several peaks (Fig. 12) as was also seen in the radio-GC chromatogram (Fig. lOB). The compolinds were identified as α-pinene (major compolind), fi-pinene, sabinene, β-myrcene, α-phellandrene, 6-pheliandrene, dihydromyrcenol (tentative), α-terpinolene (tentative) and α-terpineol (tentative).

This shows that SOSV is not a sesqliiterpene synthase as is claimed for a fragment nlicleic acid isolated by Nam et al (Plant Mol Biol, 39: 1999-2002, 1999) and Marty (EMBL Database, Accession nlimber AJ001452), blit a monoterpene synthase, viz. an α-pinene synthase. Nam et al and Marty had isolated jlist a fragment of the cDNA and for example missed the 5'-side. Hence, the alithors were also not able to flinctionally express the protein and identified it wrongly as a sesqliiterpene synthase. The GC-MS chromatograms of the inclibations of the H64MliT protein with [3H]-GPP or PH]-FPP show the presence of one terpene prodlict for each slibstrate and comparison of the retention times and mass spectra with alithentic standards confirmed that from PHJ-OPP hnalool was prodliced (Fig. 13) and from PHJ-FPF trans-nerolidol (Fig. 14). Analysis lising enantioselective collimns showed that both hnalool and nerolidol were of the S configliration, so (3S)-(E-nerolidol and S-linalool. Characterisation. The H64N0RS encoded and his-tag plirified protein was shown to have an optimlim pH of arolind 7 for both GPP and FPP/For both slibstrates there was no preference for Mn-1 (at 1 mM) or Mg2+ (at 10 mM) and therefore a combination of the two was rolitinely lised. The affinity of the enzyme for the two slibstrates strongly differed. The Ivm for FPP was 3.2 ^.M which is in the expected range for sesqliiterpene synthases. However, for GPP the Km was >50 jiM which is highly linlislial. However, the apparent Vraax for GPP was mlich higher than for FPP.
Example 7. Analysis of targeting
We lised transient expression assays lising the Green Flliorescent Protein (GFP) to identify the sliβ-celililar localization of the proteins encoded by the different nlicleic acid fragments described in this invention (Fig. 15). We first constrlicted 13 different constrlicts which flised in-frame the 5'-end parts of the different genes (H64N0RL, H64NORS, H64TAR4, H64VES, SOSV) to the GFP gene (Fig. 16). Different regions of the 5'-ends were lised part of them incllided a portion firom the protein itself (lip to the MID motif). Expression in plants was driven by the 35S caliliflower mosaic virlis promoter. Plasmid DNA firom constrlicts was lised to transform tobacco protoplasts. After transformation the protoplasts were inclibated for 24 hr at 28°C in the dark and thereafter lised for the analysis of GFP transient expression and slibcelllilar

localization lising confocal laser scanning microscopy. The reslilts demonstrated that the 5'-ends of both H64TAR4 and H64VES encode a targeting signal (Fig. 15). The protein encoded by H64TAR4 is targeted to the plastids (e.g. chloroplasts) and mitochondria while the H64VES protein is targeted to the plastids (e.g. chloroplast3). H64NORL and H64NORS, which are most active in the ripe cliltivated strawberry, are targeted to the cytosol. SOSV is also targeted to the cytosol, in-contrast to all monoterpene synthases described to date which are plastid localized. Thlis, according to this experiment for monoterpene synthases the cytosol and not only the plastids arc a possible location and in the cytosol there are high levels of GPP to synthesize the monoterpenes. For sesqliiterpene synthases normally reported to be localized in the cytosol other sliβ-celllilar localization may be possible slich as in the mitochondria and chloroplasts and they may lise FPP in these compartments and prodlice high levels of the sesqliiterpene. We also demonstrated by the same method that the different targeting signals of the terpene synthases colild be easily swapped by the lise of site-directed mlitagenesis. For example the plastidic targeting signal encoded by the H64VES N-terminal part colild be modified to dlial targeting to mitochondria and chloroplasts by a change in 2 amino acid residlies (Tryptophan-W6 changed to Arginine-R6 and deletion of Isolelicine-116).
Example 8. Effects of neroiidol on Agrobacterilim tlimefaciens FPP, the preclirsor for sesqliiterpene biosynthesis is a most common metabolite and exists in every living organism. Thlis, the expression of a protein encoding a neroiidol synthase will resvdt in the conversion of endogenolis FPP to neroiidol in most living organisms. We constrlicted a binary vector (plasmid lised for the transformation of plants celis, which lacks the virlilent genes present on the Ti plasmid of the virlilent strain oi Agrobacterilim tlimefaciens) containing the H64NORS gene flanked by a 35S Ca]\W promoter (5'-end) and a Nopaline Synthase (NOS) terminator (3'-end) and lised it to transform 2 different strains of Agrobacterilim. In both cases no colonies were obtained after plating the transformation reaction on Lmia Broth (LB) medilim containing 50 mg/1 kanamycin and Rifampicin. Thlis, the H64 NORS gene was expressed in Agrobacterilim and the protein encoded by it converted the bacterial endogenolis FPP to neroiidol, which is highly toxic to the Agrobacterilim cells, and

therefore no transformants were obtained. Thlis, transgenic plants expressing a nerolidol synthase will have an anti-microbial effect and colild be lised for the protection against Agrobacterilitn crown-gall disease. In order to be able to introdlice a plasmid containing slich a terpene synthase having toxic effects on the bacteria one can introdlice one or more introns into the coding seqlience of the gene. These introns can not be spliced by the bacteria and hence no flinctional protein is formed by the micro-organism. In the plant, the normal elikaryotic splicing process will lead to a flinctional protein. The introdliction of sliitable, organ-specific and/or indlicible promoters in the appropriate constrlict wlil allow the directed expression of hnalool and/or nerolidol at the appropriate site to control crown-gali disease in plants slich as frliits, rose, etc. Also, slow release formlilations or other compositions containing linalool and/or nerolidol may be liseflil to control crown-gall disease.
Example 9. Effects of linalool and nerolidol on spore germination, lesion growth and sporlilation of Phytophthora infestans, Flisarilim spp. and Botrytis spp.
Comparison of effects of farnesol and linalool on mycelilim growth of Phytophthora infestans on growth medilim
Farnesol and linalool were tested in two concentrations (2% and 0.2% (v/v)) throligh the addition to PKch medilim in 6 well plates (3 ml per well). One 6 well plate per compolind was lised with two different concentrations in triplicate. All welis were inoclilated with a pllig ol Phytophthora infestans mycelilim (isolate VK98014, 1 month old) and inclibated at 20°C. On day 3, 5 and 7 the radial growth of the mycelilim was measlired. An overview of the reslilts is given in Figlire 17. The mycelilim growth oi Phytophthora infestans was inhibited completely 3, 5 and 7 days after the experiment by both the high and the low concentrations of linalool. Farnesol reslilted in a partial inhibition of mycelilim growth at both the high and the low concentration. The experiment demonstrates that linalool is more active than farnesol for the inhibition of mycelilim growth of Phytophthora infestans.

Comparison of the effects of linalool and nerolidol on mycelilim growth of Phytophthora infestans alone and in combination on growth medilim Linalool and nerolidol were tested in three concentrations (0,2%, 0.02% and 0.002% (v/v)) throligh the addition to Plich medilim in 6 well plates (3 ml per well). One 6 well plate per compolind was lised with two different concentrations in triplicate. To stlidy whether the compolinds acted directly or throligh the vapolir phase in one plate mycelia were grown on control medilim with the compolinds (0.2%) added to the medilim in the adjacent wells. Free exchange of the compolinds throligh the vapolir phase was possible this way. All wells were inoclilated with a pllig of Phytophthora infestans mycelilim (isolate VK98014, 1 week old) and inclibated at 20°C. On day 3 and 5 the radial growth of the mycelilim was measlired. The reslilts are shown in Figlires 18-21.
Figlire 18 shows that linalool is active even at the lowest concentration of 0.002% (=20 ppra). Remarkably, the effects of linalool are eqlially effective throligh the vapolir phase as throligh the medilim. Apparently this monoterpene is so volatile that the active concentrations in the vapolir and medilim phase are similar. This high activity in the vapolir phase makes linalool an attractive compolind for the protection of stored prodlicts against micro-organisms e.g. the protection of potato to Phytophthora, Phoma, and Flisarilim.
Figlire 19 shows that nerohdol is slightly more effective than linalool (compare Figlire 18) in inhibiting Phytophthora infestans mycelilim growth and that it is a strong inhibitor of mycelilim growth even at the lowest concentration of 20 ppm. In contrast to linalool the effects throligh the vapolir phase are negligible. This can be explained by the fact that the sesqliiterpene nerolidol is mlich less volatile than the monoterpene linalool.
Figlires 20 and 21 show that the action of linalool and nerolidol is stronger in combination than when taken alone. This sliggests that the simliltaneolis prodliction of these compolinds in plants colild reslilt in more effective flingal control compared to a sitliation when only one of the tw o compolinds is present.

The effect of nerolidol infiltrated in potato leaves on the germination of Phytophthora infestans spores, lesion formation and sporlilation.
Leaves of potato cliltivar Bintje wei'e vaclilim-infiltrated with a 0.05% (v/v) sollition of nerolidol. This was done by placing 6 leaflets at a time in a 50 ml blliecap with the nerolidol sollition or a water control. The tlibes were placed for 15-30 min linder vaclilim which was then sliddenly released. Good infiltration was visible by the dark green color of the leaves. The leaves had gained abolit 25% weight this way so that the actlial concentration in the leaves was in the range of ca. 0.0125%. The leaves were placed on water agar (1.5%) and inoclilated with 250-500 spores of the 4 different Phytophthora infestans isolates (race-0, IPO-c, 428-2, VK98014). The leaves were inclibated one night in the dark at 15-°C and then moved to normal lighting conditions (15'C, 16h light, 8h dark). After 7 days the leaves were scored for the formation of lesions and sporlilation. The reslilts demonstrate that also when infiltrated in potato leaves nerolidol strongly inhibits mycelilira growth, lesion formation and sporlilation at a low concentration. The effects appear to be not race-specific blit eqlially affecting the folir different isolates showing that also nerolidol provides broad resistance against this flinglis.
Effects of linalool and nerolidol on Flisarilim and Botrytis mycelial growth Flisarilim. Nerolidol and linalool were tested in a range of concentrations (10-5000 ppm) alone and in combination throligh the addition to Plich medilim in 6 well plates (3 ml per well). In the case of comparing the application of nerolidol and linalool alone to the combination of the two compounds, 100 ppm of the single compolind was for example compared to 50 + 50 ppm of the two compolinds together. One 6 well plate per compolind was lised with two different concentrations in triplicate. All wells were inoclilated with a mycelilim pllig of Flisarilim graminearlim, Flisarilim clilmorlini or F.verticillioides strain MRC826 and inclibated at 20°C. Each day the radial growth of the mycelilim was measlired. The reslilts of day 7 are given in Figlire 22. The mycelimn growth of all Flisarilim spp, was inhibited at concentrations above 10 ppm. At low concentrations nerolidol was slightly more effective than linalool in the case F. graminearlim and MRC826, At very high concentrations linalool was more effective.

The combined lise of nerolidol and linalool is at least as effective as either individlial compolind and appears to provide a more roblist inhibition against all Flisarilim species.
Botrytis. Nerolidol and linalool were tested in a range of concentrations (10-5000 ppm) alone and in combination throligh the addition to Plich medilim in 6 well plates (3 ml per well). In the case of comparing the application of nerolidol and linalool alone to the combination of the two compolinds, 100 ppm of the single compolind was for example compared to 50 + 50 ppm of the two compolinds together. One 6 well plate per compolind was lised with two different concentrations in triplicate. All welis were inoclilated with a mycelilim pllig of Botrytis cinerea isolated from grape and strawberry and inclibated at 20°C. Each day the radial growth of the mycelilim was measlired. The reslilts of day 7 are given in Figlire 23. The mycelilim growth of all Botiytis isolates was inhibited at concentrations above 10 ppra. At low concentrations nerolidol was more effective than linalool. At very high concentrations linalool or the combination of linalool and nerolidol was most effective. The combined lise of nerolidol and linalool is at least as effective as either individlial compolind and appears to provide the most roblist inhibition against all Botrytis isolates.
Effects of linalool and nerolidol on Flisarilim spore germination. A spectrophotometric assay was lised to monitor the onset of germination of Flisarilim verticillioides spores (isolates: ITEM2282, MRC3235. MRC826, MRC826-2) in sollition. The spores were dillited to a concentration of 10 spores/ml in Czapek Dox medilim and mixed with 8-4000 ppm of linalool or nerolidol. Linalool did not affect the germination of the spores at ali (Figlire 24B). Nerolidol, however, showed strong inhibition of germination at concentrations above 250 ppm (Figlire 24A). This sliggests that nerolidol provides an additional mode of control of Flisarilim verticillioides at the level of spore germination in comparison to linalool and that for the most effective control at all stages of flingal development of Flisarilim spp a combined lise of nerolidol and linalool is most appropriate.

Example 10. Transformation and characterization of Arabidopsis, potato, tomato and petlinia
Preparation of constrlicts for the transformation of plants.
Constrlicts with the H64 cDNAs are prepared for the transformation of plants in
order to yield plants that will prodlice linalool and/or nerolidol in variliolis
compartments:
The cDNAs are placed linder the control of either the 35S promoter or the Rlibisco
promoter, both separately and in combination in order to obtain plants prodlicing
linalool or nerolidol alone or in combination. It is also contemplated that for some
plirposes glycosylation or deglycosylation of the terpene-alcohol is reqliired for the
mode of action against flingi or insects. For this reason also constrlicts are made
containing glycosyl-transferases or glycosidases in conjlinction with the Hnalool
and/or nerolidol synthase cDNAs.
Constrliction of binary vectors.
The appropriate seqliences were ligated into a pFlaplO vector. The ligation prodlict was transformed to E.coli DH5a competent cells, and transformed colonies were grown 0/N at 37**C and 250 rpm. The expression cassette was removed from the reslilting vector by lising Pad and Ascl restriction enzymes (NEB, England) and ligated into the binary vector pBINPLliS, containing a kanamycin resistance selection marker (nptll), after digestion with Pad and Ascl. Colonies were checked after transformation by back-transformation to E. coli DH5a competent cells.
Transformation of Arabidopsis
We lised the floral-dip transformation method to transform Arabidopsis plants ecotype Collimbia according to Marsh-Martinez et al. (2002). After collecting the seeds they were let to dry for several days and then sown on MS medilim containing

50 mg/1 kanamycin and 400 mg/1 cefotaxime. Green shoots, 1 cm m size were transferred to the green holise and grown to matlirity.
Transformation of potato
On day 1 an Agrobacterilim twnefaciens cliltlire of AGLO containing a BINPLliS derived binary vector was started in 50 ml Lβ-medilim containing 50 mgA kanamycin and shaken for 2 days at 28 oC. On day 2 internodes from an in vitro cliltlire of the potato cliltivar Desiree were clit into 0.5-1 cm pieces and placed on R3B medilim (30 g/l slicrose, 4.7 gA Mlirashige and Skoog salts, pH 5.8 (KOH), 8 gfl plirified agar, 2 mg/1 NAA and 1 mgA BAP) which was covered with 2 sterile filterpapers that had previolisly been soaked in 2 ml PACM medilim (30 g/1 slicrose, 4.7 gA Mlirashige and Skoog salts, 2 gA casein hydrolysate, pH 6.5 (KOH), 1 mg/ml 2,4-D and 0.5 mgA kinetine). The dishes were taped with parafilm and inclibated overnight at 24 oC linder a regime of 16 h light. At day 3 the A. tlimefaciens cliltlire was polired in a sterile petridish containing the explants. After 5-10 min explants are removed from the cliltlii'e, placed on a sterile filter paper to remove excess Agrobacteria and placed back on the R3B medilim containing dishes after first removing the top filter paper (leaving one behind). Dishes with explants were flirther inclibated at 24 oC and 16 h Light lintil day 5, when the explants were transferred to dishes containing ZCMv medilim (20 gA slicrose, 4.7 gA Mlirashige and Skoog salts. pH 5.8 (KOH), 8 gA plirified agar, 1 mgA zeatine, 200 mgA vancomycin, 100 mgA kanamycin, 200 mgA claforan). On day 19 and slibseqliently every 3-4 weeks explants were transferred to new ZCVK medilim. When shoots appeared shoots were transferred to Mlirashige and Skoog medilim containing 20% slicrose (MS20). After rooting plants were transferred to the green holise
Petlinia transformation.
Leaf clittings of Petlinia W115 were transformed with Agrobacterilim tlimefaciens strain LBA4404 lising a standard plant transformation protocol (Liicker et al., The Plant Jolirnal 27: 315-324, 2001). As a control leaf clittings were also transformed with LBA4404 containing the pBINPLliS vector. Flirthermore some non-transformed leaf clittings were carried throligh the regeneration process. Rooting plants, arising

from the Agrohacterilim transformation were tested with PCR for the presence of the respective gene constrlict. Positive plants were transferred to the greenholise. All ransgenic plants were phenotypically normal and showed a normal development compared with non-transformed control plants, which had gone throligh the same regeneration process.
Tomato transformation.
The tomato cliltivar Micro-Tom' {Lycopersicon flavolir) was lised (Scott and Harbaligh, 1989). The plants were grown from seeds provided by a seed company (Beekenkamp seed, Holland). Micro-Tom seeds were first sterilised. A rinse in 70% ethanol followed by a two holir bleaching in 1.5% HC104. After bleaching, the seeds were qliickly rinsed in water twice and then washed in water for ten and sixty minlites. After sterilisation, seeds were sowed in pots, containing 80ml vermiclilite and 70ml of germination medilim containing 4.4gA MS salts with vitamins and 0.5% slicrose (pH 5.8).
After 7 days of growth in a cliltlire room (25°C), covered with 2 folds of filter paper, the cotyledons were clit linder water near the petiole and the tip with a rolling action of the scalpel, to minimize damage. The explants were placed on their backs on filter paper on feederlayers to inclibate overnight in the cliltlire room (25°C). covered with 4 folds of filter paper, linder low light conditions. After inclibation, the explants wex-e immersed in the Agrobacterilim slispension for 20 minlites. After immersion, the explants were placed back on feederlayers for co-cliltivation, following a rinse in a sollition containing 400mg/l carbenicillin and 100mg/1 tricarcillin. The explants were placed in calllis indlicing mediimi (4.4gA MS salts with Nitsch vitamins, 3% slicrose, 0.8% plirified agar (Oxoid), pH 6.0, 2mg/l zeatin, 400mgA carbenicillin, 100mg/1 tricarcillin, 100mgA kanamycin). Tlie plates were covered with 2 folds of filter paper and set to grow in a cliltlire room (25°C) linder low light conditions for 3 weeks. Formed calllis was transferred to shoot indlicing medilim (as calllis indlicing medilim, blit with lmg/1 zeatin, 200mg/l carbenicillin, 50mgA tricarcillin). These plates were set to grow linder the same conditions as the calllis-indlicing plates. Shoots formed were transferred to rooting medilim in pots (4.4g/l MS salts with Nitsch vitamins, 3% slicrose, 0.5% agargel (Sigma), pH 6.0, 0.25mg/l IBA,

50mgA kanamycin, 400mgA carbenicillin. The growing conditions remained the same. Fliliγ-grown plants were slibseqliently transferred to the greenholise.
Analysis of the transgenic plants with capillary gas chromatographγ- mass spectrometry (GC-MS).
The tisslies to be analyzed were collected in the greenholise and frozen in liqliid nitrogen. In general, 200mg frozen material was homogenized and transferred to a mortar containing I.5mL 5M CaCl2 and a small amolint of plirified sea sand. These tisslies were mixed with 0.75 mL 5M CaCb. The material was grolind rapidly and thorolighly with a pestle, inhibiting enzymatic reactions. 0.75 mL of the material was introdliced into a 1.8 mL GC vial containing a small magnetic stirrer. The vial was then closed with an alliminlim cap with a PTFE/Blitylrlibber septlim. Slibseqliently the vial was placed in a 50°C waterbath and preheated for 20 minlites while stirring. The headspace sampled dliring 30 minlites with a 100μ PDMS SPME fiber (Slipelco, Belfonte PA USA).
GC-MS analysis was performed lising a Fisons 8060 gas chromatograph directly colipled to a MD 800 mass spectrophotometer (Interscience, Breda, the Netherlands). A HP-5 collimn (50 ra x 0.32 mm, film thickness 1.05μm) was lised with He (37 kPa) as carrier gas. GC oven temperatlire was programmed as follows: 2 min 80°C, ramp to 250°C at 8° min'1 and 5 min 250°C. Mass spectra in the electron impact mode were generated at 70 eV. The compolinds were identified by comparison of GC retention indices and mass spectra with those of alithentic reference compolinds. Injection was performed by thermal desorption of the SPME fiber in the injector at 250°C dliring 1 min lising the splitless injection mode with the split valve being opened after 60 sec. Alternatively, volatiles were trapped on cartridges containing Tenax, ellited lising pentane/ether and analysed lising GC-MS essentially as described by Boliwmeester et al (1998). Transgenic Arabidopsis plants expressing H64TAR, for example, prodliced large amounts of linalool and smaller amolints of nerolidol (Figlire 25). Transgenic potato lines also prodliced slibstantial amolints of linalool, blit also the hydroxγ-derivative 8-hydroxylinalool (Figure 26). Interestingly, the native linalool of potato, which can also be detected, had a different stereochemistry as the transgenic linalool (Fig. 26), which allowed a clear distinction between native and transgenic prodlict.

Becalise it was slispected that in some of the plant species these compolinds were present in a bolind form, leaf material of Petlinia (transgenic and control samples) was harvested and frozen in liqliid nitrogen, and grolind to a fine powder in a cooled mortar and pestle. In total 60 mg of the powdered leaf material was transferred to 0.5 ml of citrate bliffer at pH 4.5, to which 140 i.li. fi-gllicosidase were added. The vial was capped and inclibated dliring 12 h at 25°C. Slibseqliently, the headspace of the vial was sampled dliring 30 minlites with 100 micron PDMS solid phase microextraction device and analysed lising GC-MS as described above. No linalool or nerolidol was detectable in samples from the lintransformed control plants, whereas in the transgenic plants both linalool and nerolidol were detected. The sample of transgenic leaf material witholit betα-gllicosidase present dliring the inclibation did not show any detectable linalool or nerolidol, indicating that ali Linalool and nerolidol is stored in the petlinia leaves in the form of its gllicoside, instead of continliolis emission as was described for linalool in the flowers of Clarkia brewerL
Identification of glycosides in transgenic plants
High-performance-liqliid-chromatography electrospraγ-ionization tandem mass spectrometry (HPLC-ESI-MS-MS) analysis of methanol extracts was performed on a triple stage qliadrlipole TSQ 7000 LC-MS-MS system with an electrospray ionization (ESI) interface (Finnigan MAT, Bremen, Germany). The temperatlire of the heated capillary was 240 °C. The ESI capillary voltage was set to 3.5 kV, reslilting in a 3.4 μA clirrent. Nitrogen served as both the sheath (70 psi) and alixiliary gas (10 L/min). Data acqliisition and evalliation were carried olit on a Personal DECstation 5000/33 (Digital Eqliipment, linterfohring, Germany) and ICIS 8.1 software (Finnigan MAT). HPLC separation was carried olit on an Elirospher 100 C-18 collimn (100 x 2 mm, 5m, Knalier, Berlin, Germany) lising a linear gradient with a flow rate of 200μL min-1 Solvent A was 5 mM ammonilim acetate in water, and solvent B was 5 mM ammonilim acetate in methanol. The gradient program was as foliows: 0-30 min 5 to 100% B.Mass spectra were acqliired in the negative mode. Prodlict ion spectra were available by collision-indliced dissociation (CID) (1.5 mTorr of Argon; -20 eV). For preparation of extracts plant leaves (3 to 7 g) were homogenized in 50 ml of 80% methanol and centrifliged (2000 g for 5 min). The residlie was washed with 50 ml of 80% methanol and the slipematants were combined. Methanol was removed in

vaclilim and the remaining aqlieolis sollition was extracted with 2 x 20 ml diethyl ether. The extract was slibjected to XAD-2 (20 cm, 1 cm inner diameter) solid phase extraction. The collimn was sliccessively washed with 50 ml water and 50 ml diethyl ether. Glycosides were elated with 80 ml methanol. The extract was concentrated in vaclilim. The residlie was dissolved in 1 ml of 50% methanol in water and analyzed by HPLC-ESI-MS-MS.
R,S-Linalyl β-D-gllicopyranoside was synthesized from R,S-hnalool and 2,3,4,6-tetrα-O-acetyl-betα-D-gllicopyxanosyl bromide according to a modified Koenigs-Knorr synthesis. For enzymatic hydrolysis an aliqliot of the methanol extract was dissolved in 2 ml of 0.2 M phosphate bliffer (pH 5.5), and 200 μl of Rohapect DSL (Rohm, Darmstadt, Germany), a pectinolytic enzyme preparation exhibiting glycosidase activity was added. After an inclibation period of 24 h at 37μ, the liberated aglycons were extracted two times by 1 ml of diethyl ether each. The combined organic layers were dried over Na2SO4 and concentrated. Mliltidimensional gas chromatography mass spectrometry (MDGC-MS) analyses were performed with tandem Fison 8160 GC connected to a Fison 8130 GC and a Fisons MD 800 qliadrlipole mass spectrometer eqliipped with Fisons MassLab software (Version 1.3). The first GC was fitted with a split injector (1:10, at 230o'C) and a flame ionization detector (at 250°C). The first GC employed a 25 m x 0.25 mm i.d. flised silica capillary collimn coated with a 0.25μm film of Dβ-Wax 20 M (J & W Scientific) for the pre-separation of the target moleclile. Separation of enantiomers was achieved with the second GC lising a 25 m x 0.25 mm i.d. flised silica capillary collimn coated with a 0.15 jxm film of 2,3-di-O-ethyl-6-O-tert. Blityl dimethylsnyl-D-cyclodextrin/PS086. The collimn in GCl was coimected by a mlilticollimn switching system (Fisons) to the collimn in GC2. The retention time of the compolind of interest was determined by GC separation while the collimn in GCl was connected to the FID. Separation of the enantiomers was achieved in the second GC after transfer of the compolind of interest from the capillary collimn in GCl to the collimn in GC2 via the switching device. The flised silica capillary collimn in GCl was maintained at 60°C then programmed to 240°C at 10°C min-1 with He gas flow at 3 ml min-1, The flised sOica capillary collimn in GC2 was maintained at 60oC (15 min) then programmed to 200°C at 2°C min-1 with He gas flow at 3 ml min-1. The compolind of interest was transferred from GCl to GC2 from

9.8 min to 10.3 min. The MS operating parameters were ionization voltage, 70 eV (electron impact ionization); ion solirce and interface temperatlire, 230oC and 240°C, respectively. Linalyl-β-D-gllicopyranoside was synthesised in order to verify the identity of the glycoside present in the transgenic petlinia tisslie transformed with S-linalool synthase. HPLC-MS/MS analysis on control and transgenic Petlinia tisslie as shown in Figlire 28, revealed that the m/z 375 ion trace (Figlire 27A) of the compolind detected in the transgenic Petlinia tisslie had the same retention time as one of the two diastereomers of (R,S)-hnalyl 13-D-gllicoside that are slightly resolved in ion trace A. Also the prodlict ion spectrlim of the synthesised reference compolind fits the spectrlim of the peak detected in the transgenic petlinia tisslie nicely (Figlire 28B). The control Petlinia tisslie ion trace m/z 375 showed only a slight elevation above backgrolind level at the retention time of the linalyl β-D-gllicoside indicating that there is also a basal level of linalyl-β-D-gllicoside present in the plant before transformation (Figlire 28A). Following Chiral phase Mliltidimensional Gas Chromatography Mass Spectrometry (MDGC-MS) analysis, after enzymatic hydrolysis of the gllicoside fraction of leaf tisslie, revealed that the transgenic Petlinia leaf contains highly enriched (S)-linalyl-6-D-gllicoside. The control plant however contains shghtly enriched (fl)-linalyl-β-D-gllicoside. Since no tisslie-specific promoter for expression was lised, the enzyme can be formed in all plant organs and will give a prodlict in all cells where GPP is present. By the action of a highly active endogenolis gllicosyltransferase of Petlinia that is able to efficiently bind the S-linalool prodliced by the transgenic plants as (S)-linalyl-β-D-gllicoside, celllilar damage is prevented. Slich a highly active glycosyltransferase was also reported in transgenic Kiwi frliit expressing stilbene synthase, that acclimlilated picied (resveratrol-gllicoside) in stead of resveratrol. Large-scale volatilisation of linalool from the transgenic plants colild be excllided, since only traces of linalool were detectable when the headspace of the transformed plants was analysed. Volatilisation only occlirred from the flowers and not from leaves. This in contrast to Arabidopsis where large amolints of linalool were emitted from the leaves (Figlire 25). Therefore we concllide that most of the linalool in Pettmia is directly bolind as a β-D-gllicoside.

Flirther analysis of potato-leaf extracts also showed the presence of gllicosides, not only of linalool itself blit also of 8-hydroxylinalool. In addition, more derivatives of linalool were folind slich as linalool-triol, inclliding the corresponding gllicoside (Table 1).
In concllision, transgenic plants expressing the inserted transgenes are shown to prodlice the expected terpenoid compolinds. Their amolints, release, oxidation to polyols, and derivatization to glycosides vary from species to species and can be infllienced by the co-expression of other seqliences (see Example 11). When these compolinds are not stored in any bolind intermediates slich as glycosides, the plants have altered olfactory characteristics.





Example 11. Effects of changes in targeting of terpenoid and GPP and FPP synthases
Example 10 shows that we are able to prodlice large amolints of monoterpenes in a range of different species, and that it is not necessary to introdlice a GPP synthase linlike was claimed in WO 0022150. However, the prodliction of sesqliiterpenes in transgenic plants proved difficlilt, as has been reported by several other alithors (Wallaart et al., Planta 212: 460-465, 2001; Hohn and Ohlrogge, Plant Physiol 97: 460-462. 1991).
Therefore, the effect of introdlicing an additional gene encoding an FPP synthase to wild-type plants or plants which already over-express a sesqliiterpene synthase on sesqliiterpene prodliction was examined lising Arabidopsis thaliana plants (ecotype Collimbia). Two genes were introdliced in two ways, either by co-transformation of two binary vectors harboring the different genes or by retransforming a plant already transformed with a single gene, and selecting on a new selectable marker (hygromycin instead of kanamycin that was lised in the fii'st gene transformation). Single genes incllided an FPP synthase from strawberry encoding a cytosolic protein (FPPS), FPP synthase with a plastid targeting signal (TARFPPS) and a germacrene A sjTithase gene (GERA, sesqliiterpene synthase) that was isolated from chicory (PCT/EP 0002130), Co-transformation was performed with the following combinations of genes:
1. FPPS and a plastid localized H64NORS (H64TAR)
2. TARFPPS and H64TAE
3. FPPS and H64NORS (cytosohc)
4. FPPS and GERA.
To prevent problems with the introdliction of the H64N0RS containing plasmid to Agrobacterilim (see Example 8) we introdliced an intron to the H 64NORS gene which can not be processed by the bacteria blit can be processed by the plant. The combination of cytosolic FPP synthase and a plastidic sesqliiterpene synthase was performed in order to check whether the pool of FPP in the plastids can be increased by transport from the cytosol to the plastids. A nlimber of selfed progeny lines of a

high linalool and nerolidol prodlicing line dlie to the introdliction ot a plastidic H64NORS (H64TAR) were also co-transformed with the FPPS and TARFPPS genes. GC-MS analysis of the reslilting trangenic plants showed that the co-expression of a cytosolic FPP synthase and cytosolic sesqliiterpene synthase greatly enhances the prodliction of the sesqliiterpenes nerolidol (combination 3) and germacrene A (combination 4). Also, overexpvession of a cytosolic FPP synthase did enhance nerolidol prodliction in case of thie plastid-targeted H64NORS (combination 1). Co-expression of an FPP synthase and H64N0RS in the plastids (combination 2) did reslilt in appreciable levels of both linalool and nerolidol. This shows that it is possible to boost (transgenic) sesqliiterpene biosynthesis in the cytosol by enhancing slibstrate availability and that it is possible to direct sesqliiterpene biosynthesis to other compartmments than the cytosol.
Example 12, Transgenic plants with improved biological control of pests
Linalool and nerolidol, and its derivative 4,8-dimethyr-l,3(E).7-nonatriene have been reported to play an important role in the attraction of predators of a variety of insect and spider mite pests by a large nlimber of crops. The seqliences described in the present invention can be lised as markers for the selection of crop species, slich as for example maize, cotton, apple, and cliclimber, and any other crops employing this indirect defense mechanism, with improved prodliction of volatile, predator attracting, compolinds in response to feeding pests. In addition, the present invention can be lised to make transgenic plants with improved signalling capacity. Hereto the DNA seqliences are placed linder the control of an indlicible promoter, slich as wolind-indlicible or specific indlicible promoters. The latter type of promoters are isolated from plants that were fed lipon by for example spider mites or insects. Spider mite indlicible promoters can for example be isolated from cliclimber or lima bean. These plant species have been shown to strongly react to spider mite feeding with the prodliction of volatile signaling compolinds (Boliwmeester et al., 1999). Slibtractive (lip- and down-reglilated) libraries are made from non-infested (control) and infested plant material lising the PCR-Select™ cDNA Slibtraction Kit (Clontech). and the expression of the cDNAs in these slibtractive libraries checked lising cDNA micro-

array technology (see for example Aharoni et al., 2000) lising mRNA from control, spider-mite infested and Jα-treated plant materials as probes for hybridisation. Reglilated cDNAs will be seqlienced, and BLASTed to hint their identity. The flill-length cDNAs of interesting, strongly reglilated genes are obtained lising the RACE PCR technology, or by screening a cDNA library. The interesting flill-length cDNAs are characterised for their expression profile and lising heterologolis expression or model plant transformation. Promoters of strongly (lip-)reglilated genes are isolated lising the Genome Walker™ kit (Clontech).
As mentioned above the DNA seqliences from the invention are placed linder the control of wolind-indlicible or the isolated sliitable (tisslie-) specific (indlicible) promoters and lised for transformation of crops in which biological control is enabled by the prodliction of indlicible volatile signaliing compolinds, slich as cliclimber, maize and cotton, lising pliblished protocols. Transformation of cliclimber is carried olit lising a protocol for gene transfer and regeneration of cliclimber as developed by a Dlitch seed company.
The altered response of the transgenic plants is determined lising jasmonic acid treated and spider mite or beet army worm infested transgenic and control plants. Volatile prodliction is determined lising headspace trapping and GC-MS analysis (Boliwmeester et al., 1999). The response of the respective predators is determined lising behavioliral stlidies lising a γ-tlibe olfactometer (e.g. Takabayashi et al., J. Chem. EcoL 20(2). 373-385,1994). The transgenic Arabidopsis from Example 10 was shown to prodlice large amovmts of linalool and smaller amolints of nerolidoL In a γ-tlibe experiment, starved predatory mites (Phytoseilillis persimilis) were shown to strongly prefer the linalool/nerolidol prodlicing plants: 70% of the tested predatory mites preferred the transgenic Arabidopsis above the wild type.
Example 13. Effects of linalool and nerolidol expression on resistance to micro-organisms
Several plant species expressing the H64NORS gene and prodlicing elevated levels of linalool and nerolidol were analyzed for resistance to microbial infections of powdery mildew and Phytophthora infestans. Clear effects were observed on leaves and frliits

showing that the in vitro data presented in Example 7 are predictive of the in vivo data in transgenic plants.
Petlinia and powdery mildew
Transformed tomato plants (control (empty vector) and transgenic homozygolis for the trait) were grown from seed in a small greenholise linder identical controlled conditions (n=30). The plants were inoclilated with powdery mildew (Erysiphe cichoracearlim) spores. After 4 weeks plants were scored for infection. The reslilts indicate that the the presence of linalool protected the plants from infection by mildew (Table 2).

Tomato and Phytophthora infestans
Green firliits were harvested from variolis homozygolis transgenic Microtom tomato lines. Earlier these lines had been characterized for linsdool content by steam destination and GC-MS. Ten different berries from each transgenic line were inoclilated by pricking the top of the frliit with a tooth pick dipped in a slispension of 10,000 sporangia/ml of Phytophthora infestans IP0428-2. After 7 days the frliits were scored for infection level (Table 3). Nearly all diseased frliits had tlirned completely grey/black jlist below the skin. Frliits were scored clean if they had no infection at ali. A strong correlation was observed between a high linalool expression level and a low percentage of diseased berries. The transgenic frliits with high linalool levels largely remained free of infection


Potato and Phytoohthora infestans
Transgenic potato lines expressing the H64NORS gene in two different constrlicts (H64NOR and H64TAR) were analyzed for prodliction of linalool and nerolidol in the headspace lising an SPME fiber and GC-MS. The H64NOR constrlict did not yield nerolidol or linalool prodliction above the backgrolind present in potato, while the H64TAR constrlict gave very high levels of linalool and low levels nerolidol in the headspace. Both sets of plants were tested for Phytophthora infestans resistance by inoclilating 5 detached leaves in 2 replicates with spore slispensions and scoring lesion area, lesion growth and sporlilation (Table 4). A very strong correlation was observed between high linalool expression levels and strongly repressed or absent lesion growth and sporlilation.


per leaf (IP0428-2, 50.000 sporangia/ml) and scored for lesion area, growth and sporlilation (vislial score on a scale of 0-4) at the indicated days post infection (dpi)
Figlire 30 combines the data of table 1 and 4. Figlires 30 A, B and C provide the correlation in lesion size, lesion growth rate, and sporlilation respectively of Phytophthora infestans isolate IPO 428-2 plotted against the content of linalool, 8-hydroxylinalool, linalooltriol. lynalylgllicoside, 8-hydroxylinalylgllicoside and linalyltriolgllicoside content of the potato transgenic lines T or TM-9, -13, -29 and a control line. The control data from table 4 on flingal growth and sporlilation were taken to be the average vallies of the H64NOR plants (Mines) with negligible increased levels of either linalool, nerolidol or derivatives. The linalool (derivative) data provided in table 1 sire known to the art to be mlich more reliable and qliantitative than the SPME data on linalool in table 4, which jlistifies their preferred

lise. Figlire 30 demonstrates a strong dose-effect correlation of the levels of linalool (derivatives) prodliced in potato to the levels of resistance. With high levels of terpene expression clearly complete resistance to Phytophthora infestans infection was obtained. Flirthermore, figlire 30 D provides the in vitro data on the sensitivity of Phytophthora infestans isolate IP0428-2 which was lised for the in planta experiments to plire hnalool in the medilim as described in Example 9. From the comparison of the in vitro data with the in planta data of Figlire 30 it is clear that the qliantities prodliced in planta are in the same range as the qliantities reqliired in vitro to affect the mycehal growth. It is not clear, however, whether the natlirally formed alcohol and gllicoside derivatives of linalool are similarly active to inhibit flingal growth as the free linderived linalool forms and may contriblite to the effect of free linalool in a major way.
Example 14. Effect of linalool and nerolidol expression on insect resistance
Arabidopsis thaliana and Myzlis persicae
A line of Arabidopsis tllixliana transformed with the H64TAR constrlict was characterized to have a single gene insertion by Solithern blot, high headspace levels of hnalool and lower headspace levels of nerolidol (Example 10). This line was selected and selfed. Selfed seeds were sown and yoling, non-flowering plants were analysed for levels of linalool in the headspace lising SPME GC-MS analysis (Example 10). Homo- and heterozygolis plants with high levels of linalool were lised in a bioassay with Myzlis persicae female adlilts to observe repellent or deterrent effects of linalool expression (Table 5), For each experiment two leaves were taken from the plant, one from a control and one from a linalool plant and embedded next to each other in gelling wateragar of a small petridish. Ten adlilt females were placed on the inside of the lid of the petridish. After preset times the nlimber of adlilts on each of the leaves was recorded. A deterrent effect was visible over time. Initially the aphids did not display any preference blit after 2 days a very significant distriblition was observed in which 62% were on control plants and 38% were on hnalool plants. This indicates that linalool and for nerohdol are potential insect deterrents or repellents in plants that can express high levels of these compolinds.

Table 5. Effect of linalool prodliction in transgenic Arabidopsis on choice of aphids.


Seqliences
Note: Nlicleic acid sequence (A); Translation of nlicleic acid seqlience of A (B) or if seqlience contains an intron: (A) Nlicleic acid seqlience inclliding intron; (B) NlicJeic acid seqlience from (A) exclliding intron; (C) Translation of nlicleic acid from (B).
INFORMATION FOR NO: lA (H64NORL)
SEQliENCE CHARACTERISTICS:
LENGTH: 1874
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE
SEQliENCE DISCRIPTION FOR NO: lA
CTAATACGACTCACTATAGGGCAAGCAGTGGTAACAACGCAGAGTACGCGGGGA
CAACTTAAGTTCTTAATTCGCAAACAAAGATCAAGAAGAGCGAAAGAAATATCAT
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC
CTTCTTTAAAGTATTCAATCCCTAAATTGCCTGCTACTTTGAGATAGCTTGCTTC
CCAGTTCTATTACTATAAAGCCGATGAACGTTGAAACCAAGCATACTAGAACTAT
GGGTGACATTTTTGTCCAACATTCTCAGAAGTGGGAACTATTGAAAACTGTCTTG
AGGAATGTAGCAGAGCTAGATGCCCTTGAAGGTTTGAATATGATCGATGCTGTTC
AAAGGCTAGGCATCGATTACAACTTTCAACGAGAAATCGACGAAATCCTGCACAA
GCAAATGAGTATTGTGTCTGCCCGTGATGATCTTCATGAGGTTGCACTTCGCTTT
CGACTACTGAGACAACATGGTTACTTCGTGCCTGAAGATGTGTTTAACAACTTCA
AGGACAGCAAAGGAACGTTCAAGCAAGTTCTGGGTGAAGACATCAAGGGATTGA
TGAGCTTATACGAAGCTTCGCAGCTAGGTACAGAAGGAGAAGATATACTTGTTGA
AGCTGAAAAGTTTAGCGGCCATCTGCTAAAGACTTCTCTGTCACATCTTGATCAT
CATCGAGTCAGAATTGTTGCAAATACATTGAGGAATCCTCATCACAAAAGCTTGG
CCCCATTCATGGCCAGGAACTTTTTCGTTACTTCTCAAGCCACCAATTCATGGTTA

AATTTGCTAAAAGAAGTAGCAAAAACAGATTTCAATATGGTCCGGTCTCTGCACC
AGAATGAAATAGTTCAAATGTCCAAATGGTGGAAGGAGCTTGGATTGGCTAAGG
AACTGAAGTTTGCAAGAGATCAACCACTGAAATGGTACATTTGGTCCATGGCATG
CCTGACAGATCCAAAGTTATCAGAGGAGAGGGTTGAGCTCACAAAACCCATCTCT
TTTGTCTATTTGATAGATGACATTTTCGATGTTTATGGAACCCTTGATGACCTCAT
TCTCTTCACAGAAGCTGTTAATCGATGGGAAATTACTGCTATAGACCACTTACCA
GACTATATGAAGATATGCTTCAAGGCTCTCTATGATATGACTAATGAATTCAGCA
GCAAGGTCTATCTGAAGCATGGATGGAACCGCTTACAATCTTTGAAAATTTCGTG
GGCGAGTCTTTGCAATGCATTTTTGGTGGAAGCAAAATGGTTCGCCTCTGGGAAG
CTGCCGAAGTCAGAAGAGTACTTGAAGAATGGCATCGTTTCTTCTGGGGTAAATG
TGGTTCTAGTCCACATGTTTTTTCTCTTGGGTCAGAACATAACCAGAAAGAGTGT
GGAGTTGTTGAATGAAACTCCAGCCATTATATCGTCCTCAGCAGCAATTCTTCGA
CTCTGGGACGATTTAGGAAGTGCAAAGGATGAGAACCAGGATGGGAACGATGGG
TCGTATGTAAGGTGCTACTTAGAGGAACATGAAGGCTGTTCCATTGAGGAGGCAC
GAGAAAAGACGATTAATATGATTTCAGATGAATGGAAGAAACTGAACAGAGAACT
GCTCTCTCCAAATCCATTTCCAGCATCATTCACAtTGGCTTCTCTTAATCTCGCAA
GAATGATCCCCTTGATGTATAGCTACGATGGCAACCAATGCCTTCCATCTCTTAA
AGAGTATATGAAACTGATGTTGTATGAGACTGTATCAATGTAATTAATAATAAGA
CTACCGGAAGTGGAGTTGAACTTCAAAGGTGGGTGGTCAAGAGAAACAAGAAGC CTAAG
INFORMATION FOR NO: IB (H64NORL)
SEQliENCE CHARACTERISTICS:
LENGTH: 519
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE
SEQliENCE DISCRIPTION FOR NO: IB

MNVETKHTRTMGDIFVQHSQIvLELLKTVLRNVAELDALEGLNMIDAVQRLG
IDYNFQREIDEILHKQMSIVSAKDDLHEVALRFRLLRQHGYFVPEDVFNNFK
DSKGTFKQVLGEDIKGLMSLYEASQLGTEGEDILA'EAEKFSGHLLKTSLSHL
DHHRVRIVANTLRNPHHKSLAPFMARNFFVTSQATNSWLNLLKEVAKTDFN
MVRSLHQNEIVQMSKVVWKELGLAKELKFARDQPLKWYIWSMACLTDPKLS
EERVELTKPISFVYLIDDIFDVYGTLDDLILFTEAVNRWEITAIDHLPDYMKIC
FKALYDMTNEFSSKVYLKHGWNPLQSLKISWASLCNAFLVEAKWFASGKLP
KSEEYLKNGIVSSGVNWLVHMFFLLGQNITRKSVELLNETPAIISSSAAILRL
WDDLGSAKDENQDGNDGSYVRCYLEEHEGCSIEEAREKTINMISDEWKKLN
RELLSPNPFPASFTLASLNLARMIPLMYSYDGNQCLPSLICEYMKLMLYETVS
M
INFORMATION FOR NO: 2A (H64NORS)
SEQliENCE CHARACTERISTICS:
LENGTH: 1631
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
liNALOOL/NEROLIDOL SYNTHASE
SEQliENCE DISCRIPTION FOR NO: 2A
ATGAACGTTGAAACCAAGCATACTAGAACTATGGGTGACATTTTTGTCCAACATT
CTCAGAAGTTGGAACTATTGAAAACTGTCTTGAGGAATGTAGCAGAGCTAGATGC
CCTTGAAGGTTTGAATATGATCGATGCTGTTCAAAGGCTAGGCATCGATTACAAC
TTTCAACGAGAAATCGACGAAATCCTGCACAAGCAAATGAGTATTGTGTCTGCCC
GTGATGATCTTCATGAGGTTGCACTTCGCTTTCGACTACTGAGACAACATGGTTA
CTTCGTGCCTGAAGATGTGTTTAACAACTTCAAGGACAGCAAAGGAACGTTCAAG
CAAGTTCTGGGTGAAGACATCAAGGGATTGATGAGCTTATACGAAGCTTCGCAGC
TAGGTACAGAAGGAGAAGATATACTTGTTGAAGCTGAAAAGTTTAGCGGCCATCT

GCTAAAGACTTCTCTGTCACATCTTGATCATCATCGAGTCAGAATTGTTGCAAATA
CATTGAGGAATCCTCATCACAAAAGCTTGGCCCCATTCATGGCCAGGAACTTTTT
CGTTACTTCTCAAGCCACCAATTCATGGTTAAATTTGCTAAAAGAAGTAGCAAAA
ACAGATTTCAATATGGTCCGGTCTCTGCACCAGAATGAAATAGTTCAAATGTCCA
AATGGTGGAAGGAGCTTGGATTGGCTAAGGAACTGAAGTTTGCAAGAGATCAAC
CACTGAAATGGTACATTTGGTCCATGGCATGCCTGACAGATCCAAAGTTATCAGA
GGAGAGGGTTGAGCTCACAAAACCCATCTCTTTTGTCTATTTGATAGATGACATT
TTCGATGTTTATGGAACCCTTGATGACCTCATTCTCTTCACAGAAGCTGTTAATCG
ATGGGAAATTACTGCTATAGACCACTTACCAGACTATATGAAGATATGCTTCAAG
GCTCTCTATGATATGACTAATGAATTCAGCAGCAAGGTCTATCTGAAGCATGGAT
GGAACCCCn^ACAATCTTTGAAAATTTCGTGGGCGAGTCTTTGCAATGCATTTTT
GGTGGAAGCAAAATGGTTCGCCTCTGGGAAGCTGCCGAAGTCAGAAGAGTACTT
GAAGAATGGCATCGTTTCTTCTGGGGTAAATGTGGTTCTAGTCCACATGTTTTTT
CTCTTGGGTCAGAACATAACCAGAAAGAGTGTGGAGTTGTTGAATGAAACTCCAG
CCATTATATCGTCCTCAGCAGCAATTCTTCGACTCTGGGACGATTTAGGAAGTGC
AAAGGATGAGAACCAGGATGGGAACGATGGGTCGTATGTAAGGTGCTACTTAGA
GGAACATGAAGGCTGTTCCATTGAGGAGGCACGAGAAAAGACGATTAATATGAT
TTCAGATGAATGGAAGAAACTGAACAGAGAACTGCTCTCTCCAAATCCATTTCCA
GCATCATTCACATTGGCTTCTCTTAATCTCGCAAGAATGATCCCCTTGATGTATAG
CTACGATGGCAACCAATGCCTTCCATCTCTTAAAGAGTATATGAAACTGATGTTG
TATGAGACTGTATCAATGTAATTAATAATAAGACTACCGGAAGTGGAGTTGAACT
TCAAAGGTGGGTGGTCAAGAGAAACAAGAAGCCTAAG
INFORMATION FOR NO: 2B (H64NORS)
SEQliENCE CHARACTERISTICS:
LENGTH: 519
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE
SEQliENCE DISCRIPTION FOR NO: 2B

MNVETIfflTRTMGDIFVQHSQKLELLKTVLRNVAELDALEGLNMIDAVQRLGIDY
NFQREIDEILHICQMSIVSARDDLHEVALRFRLLRQHGYFVPEDVFr^NFKDSKGTF
KQVLGEDIKGLMSLYEASQLGTEGEDILVEAEICFSGHLLKTSLSHLDHHRVRIVA
NTLRNPHHKSLAPFMARNFFVTSQATNSWLNLLKEVAICTDFNMVRSLHQNEIVQ
MSKWWKELGLAICELKFARDQPLKWYIWSMACLTDPIOSEERVELTKPISFVYLID
DIFDVYGTLDDLILFTEAVNRWEITAIDHLPDYMiaCFKALYDMTNEFSSKVYLKH
GWNPLQSLiaSWASLCNAFLVEAKWFASGKLPKSEEYLKNGIVSSGVNWLVHMF
FLLGQNITRKSVELLNETPAIISSSAAILRLWDDLGSAKDENQDGNDGSYVRCYLE
EHEGCSIEEAREICTIlSnVIISDEWKICLNRELLSPNPFPASFTLASLNLARMIPLMYSY
DGNQCLPSLKEYMKLMLYETVSM
INFORMATION FOR NO: 3A (H64MliT)
SEQliENCE CHARACTERISTICS:
LENGTH: 1874
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CLXTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE
SEQliENCE DISCRIPTION FOR NO: 3A
CTAATACGACTCACTATAGGGCAAGCAGTGGTAACAACGCAGAGTACGCGGGGA
CAACTTAAGTTCTTAATTCGCAAACAAAGATCAAGAAGAGCGAAAGAAATATCAT
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC
CTTCTTTAAAGTATTCAATCCTCAAATTGCCTGCTACTTTCTAGATAGCTTGCTTC
CCAGTTCTATTACTATAAAGCCGATGAACGTTGAAACCAAGCATACTAGAACTAT
GGGTGACATTTTTGTCCAACATTCTCAGAAGTTGGAACTATTGAAAACTGTCTTG
AGGAATGTAGCAGAGCTAGATGCCCTTGAAGGTTTGAATATGATCGATGCTGTTC
AAAGGCTAGGCATCGATTACAACTTTCAACGAGAAATCGACGAAATCCTGCACAA
GCAAATGAGTATTGTGTCTGCCCGTGATGATCTTCATGAGGTTGCACTTCGCTTT
CGACTACTGAGACAACATGGTTACTTCGTGCCTGAAGATGTGTTTAACAACTTCA

AGGACAGCAAAGGAACGTTCAAGCAAGTTCTGGGTGAAGACATCAAGGGATTGA
TGAGCTTATACGAAGCTTCGCAGCTAGGTACAGAAGGAGAAGATATACTTGTTGA
AGCTGAAAAGTTTAGCGGCCATCTGCTAAAGACTTCTCTGTCACATCTTGATCAT
CATCGAGTCAGAATTGTTGCAAATACATTGAGGAATCCTCATCACAAAAGCTTGG
CCCCATTCATGGCCAGGAACTTTTTCGTTACTTCTCAAGCCACCAATTCATGGTTA
AATTTGCTAAAAGAAGTAGCAAAAACAGATTTCAATATGGTCCGGTCTCTGCACC
AGAATGAAATAGTTCAAATGTCCAAATGGTGGAAGGAGCTTGGATTGGCTAAGG
AACTGAAGTTTGCAAGAGATCAACCACTGAAATGGTACATTTGGTCCATGGCATG
CCTGACAGATCCAAAGTTATCAGAGGAGAGGGTTGAGCTCACAAAACCCATCTCT
TTTGTCTATTTGATAGATGACATTTTCGATGTTTATGGAACCCTTGATGACCTCAT
TCTCTTCACAGAAGCTGTTAATCGATGGGAAATTACTGCTATAGACCACTTACCA
GACTATATGAAGATATGCTTCAAGGCTCTCTATGATATGACTAATGAATTCAGCA
GCAAGGTCTATCTGAAGCATGGATGGAACCCCTTACAATCTTTGAAAATTTCGTG
GGCGAGTCTTTGCAATGCATTTTTGGTGGAAGCAAAATGGTTCGCCTCTGGGAAG
CTGCCGAAGTCAGAAGAGTACTTGAAGAATGGCATCGTTTCTTCTGGGGTAAATG
TGGTTCTAGTCCACATGTTTTTTCTCTTGGGTCAGAACATAACCAGAAAGAGTGT
GGAGTTGTTGAATGAAACTCCAGCCATTATATCGTCCTCAGCAGCAATTCTTCGA
CTCTGGGACGATTTAGGAAGTGCAAAGGATGAGAACCAGGATGGGAACGATGGG
TCGTATGTAAGGTGCTACTTAGAGGAACATGAAGGCTGTTCCATTGAGGAGGCAC
GAGAAAAGACGATTAATATGATTTCAGATGAATGGAAGAAACTGAACAGAGAACT
GCTCTCTCCAAATCCATTTCCAGCATCATTCACATTGGCTTCTCTTAATCTCGCAA
GAATGATCCCCTTGATGTATAGCTACGATGGCAACCAATGCCTTCCATCTCTTAA
AGAGTATATGAAACTGATGTTGTATGAGACTGTATCAATGTAATTAATAATAAGA
CTACCGGAAGTGGAGTTGAACTTCAAAGGTGGGTGGTCAAGAGAAACAAGAAGC
CTAAG

INFORMATION FOR NO: 3B (H64MliT)
SEQliENCE CHARACTERISTICS:
LENGTH: 552
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE
SEQliENCE DISCRIPTION FOR NO: 3B
MASSSRAFFKVFNPQIACYFLDSLLPSSITIKPMNVETKHTRTMGDIFVQHSQICLE
LLKTVLRNVAELDALEGLNMIDAVQRLGIDYNFQREIDEILHKQMSrVSARDDLHE
VALRFRLLRQHGYFVPEDVFNNFKDSKGTFKQVLGEDIKGLMSLYEASQLGTEGE
DILVEAEI^SGHLLKTSLSHLDHHRVRIVANTLRNPHHKSLAPFMARNFFVTSQA
TNSWLNLlia:VAiaT)FNMVRSIJIQNEIVQMSKWWI YTWSMACLTDPKLSEERVELTKPISFVYLIDDIFDVYGTLDDLILFTEA\^^JRW'EITAI
DHLPDYMKICnvALYDMTNEFSSKVYlivHGWNPLQSLKISWASLCNAFLVEAKVV
FASGIOPKSEEYLIvNGIVSSGVNWLVHMFFLLGQNITRKSVELLNETPAIISSSAA
ILRLWDDLGSAKDENQDGNDGSY\TlCYLEEHEGCSIEEAREKTIN'NMSDEWKKLN
RELLPNPFPASFTLASLNIARMVnPLMYSYDGNQCLPSLKEYMKLMLYETVSM
INFORMATION FOR NO: 4A (H64VES)
SEQliENCE CHARACTERISTICS:
LENGTH: 1894
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: WILD STRAWBERRY LINALOOL/NEROLIDOL
SYNTHASE
SEQliENCE DISCRIPTION FOR NO: 4A

AAGCAGTGGTAACAACGCAGAGTACGCGGGGACAACTGAAGTTCTTAATTCGCA
AACAAAGATCAAGAAGAGCGAAAGAAGTATCATCTCCCGCCTTAGGTGCTGATCA
TAGATCAGATGGCATCGTCTTCTTGGGCCTTCTTTAAAGTATTCAATCCCCAAATT
GCTCCAAAAAGTATCTCACATATTGGCCAGTCTGACCTCATGCAGCTTACACATA
AGAAGCAGCTGCCTACTTTTCAAAGACGGGGCATTGCCGAAGATAGCTTGCTTCC
CAGTTCTACTACTCCCATAAAGCCGATGCACGTTGAAACCAAGCATACTAGAACT
ATGGGTGACATTTTTGTCCAACATTCTCAGAAGTTGGAACTATTCAGAAATGTCTT
GAGGAATGCAGCAGAGCTAGATGCCCTTGAAGGTTTGAATATGATCGATGCCGTT
CAAAGGCTAGGCATCGATTACCACTTTCAACGAGAAATCGACGAAATTCTGCACA
AGCAAATGGGTATTGTATCTGCCTGTGATGATCTTTATGAGGTTGCACTTCGTTT
TCGACTACTGAGACAACATGGTTACTTCGTGCCTGAAGATGTGTTTAACAACTTC
AAGGACAGCAAAGGAACTTTCAAGCAAGTTCTGGGTGAAGACATCAAGGGATTG
ATGAGCTTATACGAAGCTTCGCAGCTAGGTACAGAAGGAGAAGATACACTTGTTG
AAGCTGAAAAGTTTAGTGGCCATCTGCTAAAGACTTCTCTGTCACATCTTGATCG
TCATCGAGCCAGAATTGTTGGAAATACATTGAGGAATCGTCATCGCAAAAGCTTG
GCCTCATTCATGGCCAGGAACTITTTCGTTACTTCTCAAGCCACCAATTCATGGTT
AAATTTGCTAAAAGAAGTAGCAAAAACAGATTTCAATATGGTCCGGTCTGTGCAC
CAGAAAGAAATAGTTCAAATTTCCAAATGGTGGAAGGAGCTTGGATTGGTTAAGG
AACTGAAGTTTGCAAGAGATCAACCACTGAAATGGTACACTTGGTCCATGGCAGG
CCTAACAGATCCAAAGTTATCAGAGGAGAGGGTTGAGCTCACAAAACCCATCTCT
TTTGTCTATTTGATAGATGACATTTrCGATGTTTATGGAACCCTTGATGACCTCAT
TCTCTTCACAGAAGCTGTTAATAGATGGGAAATTACTGCTATAGACCACTTACCA
GACTATATGAAGATATGGTTCAAGGCTCTCTATGATATGACTAATGAATTCAGCT
GCAAGGTCTATCAGAAGCATGGATGGAACCCCTTACGATCTTTGAAAA'nTCGTG
GGCGAGTCTTTGCAATGCGTTTTTGGTGGAAGCAAAATGGTTCGCATCTGGGCA
GCTGCCGAAGTCAGAAGAGTACTTGAAGAACGGCATCGTTTCTTCTGGGGTAAAT
GTGGGTCTAGTCCACATGTTTTTTCTCTTGGGTCAGAACATAACCAGAAAGAGTG
TGGAGTTGTTGAATGAAACTCCAGCCATGATATCGTCCTCAGCAGCAATTCTTCG
ACTCTGGGACGATTTAGGCAGTGCAAAGGATGAGAACCAGGATGGGAACGATGG
GTCGTATGTAAGGTGCTACTTAGAGGAACATGAAGGCTGTTCCATTGAGGAGGC
ACGAGAAAAGACGATTAATATGATTTCAGATGAATGGAAGAAACTGAACAGAGAA

CTGCTCTCTCCAAATCCATTTCCAGCAACATTCACATCGGCTTCTCTTAATCTCGC AAGAATGATCCCCTTGATGTATAGCTACGATGGCAACCAATCCCTTCCATCTCTT
AAAGAGTATATGAAACTGATGTTGTATGAGACTGTATCAATGTAAITGATAATAA GACTGCTGGAAGTGGAGTTGAACA
INFORMATION FOR NO: 4B (H64VES)
SEQliENCE CHARACTERISTICS.
LENGTH: 580
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: WILD STRAWBERRY LINALOOL/NEROLIDOL
SYNTHASE
SEQliENCE DISCRIPTION FOR NO: 4B
MASSSWAFFKVFNPQLAPKSISHIGQ8DLMQLTHKKQLPTFQRRGliEDSLLPSST
TPIKPMHVETKHTRTMGDIFV'QHSQKLELFRNVLRNAAELDALEGLNiVLAAVQRL
GIDYHFQREIDEILHKQMGIVSACDDLYEVALRFRLLRQHGYFVPEDVFNNFKDS
KGTFKQVLGEDIKGLMSLYEASQLGTEGEDTLVEAEKFSGHLLKTSLSHLDRHRA
RrV-GNTlilNPHRKSlJ\SFMARNFFVTSQATNSWLJ^LKEVAKTDFNMVRSVHQK
EIVQISKWWKEI/5LVKELKFARDQPLKWYTWSMAGLTDPKLSEERVELTKPISFV
YLTODIFDVYGTLDDliLFTEAVNRWEITAIDHLPDYMiaCFKALYDMTNEFSCKV
YQKHGWNPliJSIJaSWASLCNAFLVEAIvWFASGQLPKSEEYLKN'GIVSSGVNVGL
VHMFFLLGQNITRKSVELLNETPAMISSSAAILRLWDDLGSAKDENQDGNDGSYV
RCYLEEHEGCSIEEAREKTINMISDEWKLANRELLSPNPFPATFTSASLNLARMIP
LMYSYDGNQSLPSLKEYMKLVILYETVSM

INFORMATION FOR NO: 5A (H64NORD1/W155)
SEQliENCE CHARACTERISTICS:
LENGTH: 333
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Incllides stop codon and an intron.
SEQliENCE DISCRIPTION FOR NO: 5A
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC
CTTCTTTAAAGTATTCAATCCTCAAATTGCCTGCTACTTTTGAGATAGCTTGCTTC
CCAGTTCTATTACTATAAAGCCGATGAACGTTGAAACCAAGCATACTAGAACTAT
GGTAAAATTCTCGGAGCTTTCTCCGAAGTACATTTCTACAAAAGGGTAGAGCTAG
CTACTAAACAATAGTTAATTGACTGTGCCTTGCTTGCAGGGTGACATTTTTGTCCA
ACATTCTCAGAAGTTGGAACTATTGAAAACTGTCTTGAGGAATGTAGCAGAGCTA
G
INFORMATION FOR NO: 5B (H64NORD1AV155)
SEQliENCE CHARACTERISTICS:
LENGTH: 240
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Stop codon modified and intron removed.
SEQliENCE DISCRIPTION FOR N0:5B
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC CTTCTTTAAAGTATTCAATCCTCAAATTGCCTGCTACTTTCTAGATAGCTTGCTTC CCAGTTCTATTACTATAAAGCCGATGAACGTTGAAACCAAGCATACTAGAACTAT GGGTGACATTTTTGTCCAACATTCTCAGAAGTTGGAACTATTGAAAACTGTCTTG AGGAATGTAGCAGAGCTAG

INFORMATION FOR NO: 5C (H64NORD1AV155)
SEQliENCE CHARACTERISTICS:
LENGTH: 68
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Stop codon modified and intron removed and fragment translated.
SEQliENCE DISCRIPTION FOR NO: 5C
MASSSRAFFKVFNPQIACYFLDSLLPSSITIKPMNVETKHTRTMGDIFVQHSQKLE LLKTVLRNVAEL
INFORMATION FOR NO: 6A (H64NORli1AVI51)
SEQliENCE CHARACTERISTICS:
LENGTH: 392
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Incllides stop codon and an intron.
SEQliENCE DISCRIPTION FOR NO: 6A
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATAGTCTTTTCGGTC
CCTCTTTAAAGTATTCAATCAAATTGCTCCAAAAATTATCTCACATGTTGGCCACT
CTAAGAAGCAGCTGCCTGCTACTTTTCAAAGATGGGGCGTTGCCGAAGATAGCTT
GCTTTCCAGTTCTAGTACTATAAAGCTGATGAACGTTGAAACCAAGCATACTAGA
ACTATGGTAAAATTCTTGGGGCTTTCTCCTACGTACATTTCTTCAATGAGGCTAGC
TAGCTACTAAACAATAGTTAATTGACTGTGCCTTACTGGCAGGATGACATTTTTGT
CCAACATTCTCGGAAGCTGGAACTACTCAGGAATGTCTTGAGGAATGTAGCAGAG
CTAG

INFORMATION FOR NO: 6B (H64NORli1AV151)
SEQliENCE CHARACTERISTICS:
LENGTH: 300
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Stop codon modified and intron removed.
SEQliENCE DISCRIPTION FOR NO: 6B
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCAAAGTCTTTTCGGTC
CCTCTTTAAAGTATTCAATCAAATTGCTCCAAAAATTATCTCACATGTTGGCCACT
CTAAGAAGCAGCTGCCTGCTACTTTTCAAAGATGGGGCGTTGCCGAAGATAGCTT
GCTTTCCAGTTCTAGTACTATAAAGCTGATGAACGTTGAAACCAAGCATACTAGA
ACTATGGATGACATTTTTGTCCAACATTCTCGGAAGCTGGAACTACTCAGGAATG
TCTTGAGGAATGTAGCAGAGCTAG
INFORMATION FOR NO: 6C (H64NORli1AV151)
SEQliENCE CHARACTERISTICS:
LENGTH: 88
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Stop codon modified and intron removed and fragment translated
SEQliENCE DISCRIPTION FOR NO: 6C
MAKSFRSLFKVFNQIAPKIISHVGHSQCQLPATFQRWGVAEDSLLSSSSTIKLMN\' ETKHTRTMDDIFVQHSRKLELLRNVLRNVAEL

INFORMATION FOR NO: 7A (H64NORli2/UP3)
SEQliENCE CHARACTERISTICS:
LENGTH: 350
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Incllides stop codon and an intron.
SEQliENCE DISCRIPTION FOR NO: 7A
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATAGTCTTTTCGGTC
CCTCTTTAAAGTATTCAATCAAATTGCTCCAAAAATTATCTCACATGTTGGCCACT
CTAAGAAGCAGCTGCCTGCTACTTTTCAAAGATGGGGCGTTGCCGAAGATAGCTT
GCTTTCCAGTTCTAGTACTATAAAGCTGATGAACGTTGAAACCGAGCATACTAGA
ACTATGGTAAAATTCTTGGGGCTTTCTCCTACGTACATTTCTTCAATGAGGCTAGC
TAGCTACTAA.CCAATAGTTAATTGACTGTGCCTTACTTGCAGGATGACATTTTGT
CCAACATTCTCGGAAGC
INFORMATION FOR NO: 7B (H64NORli2A/UP3)
SEQliENCE CHARACTERISTICS:
LENGTH: 258
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Stop codon modified and intron removed.
SEQliENCE DISCRIPTION FOR NO: 7B
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCAAAGTCTTTTCGGTC CCTCTTTAAAGTATTCAATCAAATTGCTCCAAAAATTATCTCACATGTTGGCCACT CTAAGAAGCAGCTGCCTGCTACTTTTCAAAGATGGGGCGTTGCCGAAGATAGCTT GCTTTCCAGTTCTAGTACTATAAAGCTGATGAACGTTGAAACCGAGCATACTAGA ACTATGGATGACATTTTTGTCCAACATTCTCG GAAGC

INFORMATION FOR NO: 7C (H64NORli2/liP3)
SEQliENCE CHARACTERISTICS:
LENGTH: 74
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Stop codon modified and intron removed and fragment translated
SEQliENCE DISCRIPTION FOR NO: 7C
MAKSFRSLFKVFNQlAPKIISHVGHSICKQLPATFQRWGVAEDSLLSSSSTIKLMNV ETEHTRTMDDIFVQHSRK
INFORi\L\TION FOR NO: SA (H64NORli3/liP16)
SEQliENCE CHARACTERISTICS:
LENGTH: 367
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Incllides an intron.
SEQliENCE DISCRIPTION FOR NO: 8A
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC
CTTCTTTAAAGTATTCAATCCTGCTCCAAAAAGCATCCCACGTATTGGCCAGTCTA
ACCTCATGCAGCTTACACATAAGAAGCAGCTGCCTACTTTTCAAAGACGGGGCAT
TGCCGAAGATAGCTTGCTTCCCAGTTCTACTACTCCCATAAAGCTGATGAACGTT
GAAACCAAGCATACTAGAACTATGGTAAAATTCTCGGAGCTTTCTCCGAAGTACA
TTTCATCAAGAGGCTAGCTATAGCTACTACACAATAGTTTGACTGTGCCTTGCTT
GCAGGGTGACATITTTGTCCAACATTGTCAGAAGTT

INFORMATION FOR NO: 8B (H64NORli3/UP16)
SEQliENCE CHARACTERISTICS:
LENGTH: 277
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Intron removed.
SEQliENCE DISCRIPTION FOR NO: 8B
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC
CTTCTTTAAAGTATTCAATCCTGCTCCAAAAAGCATCCCACGTATTGGCCAGTCTA
ACCTCATGCAGCTTACACATAAGAAGCAGCTGCCTACTTTTCAAAGACGGGGCAT
TGCCGAAGATAGCTTGCTTCCCAGTTCTACTACTCCCATAAAGCTGATGAACGTT
GAAACCAAGCATACTAGAACTATGGGTGACATTTTTGTCCAACATTGTCAGAAGT
T
INFORMATION FOR NO: 8C (H64NORU3/UP16)
SEQliENCE CHARACTERISTICS:
LENGTH: 80
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Intron removed and fragment translated
SEQliENCE DISCRIPTION FOR NO: 8C
MASSSRAFFKVFNPAPKSIPRIGQSNLMQLTHKKQLPTFQRRGIAEDSLLPSSTTPI KLMNVETKHTRTMGDIFVQHCQK

INFORMATION FOR NO: 9A (H64NORli4/liP1)
SEQliENCE CHARACTERISTICS:
LENGTH: 357
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Incllides an intron.
SEQliENCE DISCRIPTION FOR NO: 9A
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC
CTTCTTTAAAGTATTCAATCCTGCTCCAAAAAGCATCCCACGTATTGGCCAGTCTA
ACCTCATGCAGCTTACACATAAGAAGCAGCTGCCTACTTTTCAAAGACGGGGCAT
TGCCGAAGATAGCTTGCTTCCCAGTTCTACTACTCCCATAVAAGCCGATGAACGTT
GAAACCAAGCATACTAG.GACTATGGTAAAATTCTCGGAGCTTTCTCCGAAGTACA
TTTCATCAAGAGGCTAGCTATAGCTACTACACACATAGTTGGACTGTGCCTTGCTT
GCAGGGTGACATTTTTGTCCAACATT
INFORMATION FOR NO: 9B (H64NORli4AJP1)
SEQliENCE CHARACTERISTICS:
LENGTH: 267
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Intron removed.
SEQliENCE DISCRIPTION FOR NO: 9B
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC
CTTCTTTAAAGTATTCAATCCTGCTCCAAAAAGCATCCCACGTATTGGCCAGTCTA
ACCTCATGCAGCTTACACATAAGAAGCAGCTGCCTACTTTTCAAAGACGGGGCAT

TGCCGAAGATAGCTTGCTTCCCAGTTCTACTACTCCCATAAAGCCGATGAACGTT GAAACCAAGCATACTAGAACTATGGGTGACATTTTTGTCCAACATT
INFORMATION FOR NO: 9C (H64NORli4yliP])
SEQliENCE CHARACTERISTICS:
LENGTH: 77
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: PCR fragment from the 5' of cliltivated strawberry
H64. Intron removed and fragment translated
SEQliENCE DISCRIPTION FOR NO: 9C
MASSSRAFFIvVFNPAPKSIPRIGQSNLMQLTHICKQLPTFQRRGIAEDSLLPSSTTPI
KPMNVETiaiTRTMGDIFVNI
INFORMATION FOR NO: lOA (SOSAAVS)
SEQliENCE CHARACTERISTICS:
LENGTH: 1672
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY OLEFIN SYNTHASE.
INCLliDES A STOP CODON.
SEQliENCE DISCRIPTION FOR NO: lOA
ATGCCTGTCCATGCTACTCCAGCAGCTGAATCCCAGATCATCTCTATGCCGGAAG
TTGTTCGGCGCACAGCAAATTTTAAACCTAGCGTTTGGGGAGATCGGTTTGCTAA
CTATGCCGAAGACATTATAACTCAAACTCAAATGCAAGAACAAGTTGAGGAGCTG
AAACAAGTGAGGAAGGAAGTATTCACTAATGCTGCTGATGATTCTTCACATCAAC
TGAAGCCAATTGATGAAATCCAGCGCCTCGGTGTGGCTTACCATTTCGAAAGCGA
AATAGATCAAGCCCTGGAACGTATACATGAGACATATCAAGATATTCATGATGGT
GGTGATCTGTACAATGTTGCTCTTCGTTTTCGGCTACTCAGGCGACATGGATATA
ATGTTTCGTGCGATGTATTCAACAAGTTCAAAGATACTAATGGTGACTACAAGAA

AAGCTTGGTCACTGATCTTTCTGGTATGCTGAGCTTTTATGAGGCGGCCCATCTG
AGGGTGCATGGAGAAAAATTACTTGAAGAGGCTCTGGTTTTTACCACCACTCATC
TCCAGTCAGCAAGTGCAAAAAGCTCITTGCTGAAAACACAAATAACTGAAGCCGT
AGAGAGACTACTAAAAACTATGGAGAGGTTAGGTGCTCGGCGTTACATGTCA.ATA
TATCAAGATGAAGCTTCATACAGTGAAAATTTACTGAAACTTGCAAAATTAGATTT
TAATGTTGTTCAGTGTTTACACAAAAAGGAACTCAGTGACATTCCCTAAGATGGT
ACAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCTCGAGATAGGATCGTGG
AGTTGTTCTTTTGGATAGCAGGAATATATTTCGAACCTGAATACGTCTTTAGGAG
ACACATTCTGACTAAACTGATTGAGATAACAACAGTAATGGATGATATGTATGAT
GCATTCGGTACATTCGAAGAACTCGTCAACTTGACTGAAGCAATTGACAGGTGGG
ATGCAAGTTGGATGGATCAACTGCCAGACTATATGCAACCATTTTATATTACACTT
CTGGATGTTATCGATGAAGTTGAAGAGGAGCTGACAAAGCAAGGAAGATCTTAC
CGAATTCACTACGCAAAAGAAATTATGAAGAATCAAGCCAGGCTCTACTTCGCTG
AGGCCAGATGGTTCCACGAAGGATGCACCCCAAAAATGGATGAGTATATGCGAG
TTGCGGCATCTTCTGTCGGTAACACCATGCTTTCCGTCGTGTCTTTAGTAGGCAT
GGGAGACATTATAACAAAATTTG.AATTCGAGTGGCTGACCAATGAGCCTAAAATC
C'lTAGAGCTTCGAATACCATATTTAGGCTTATGGATGACATTGCTGGGTACAAGT
TTGAGAAAGAGAGAGGGCATGTTGCTTCAAGTATTGATTGCTACATGAATGAATA
CGGGGTTTCAGAGCAAGAGACAATTGATATCTTCAACAAACGAATTGTGGATTCG
TGGAAGGATATAAACGAAGAGTTTCTGAGACCCACTGCTGCTCCAGTCCCTGTGC
TTAATCGTGTTCTTAACCTAACCCGAGTGGTTGATCTGCTTTACAAAAGGGGAGA
TGCCTTCACGCATGTCGGAAAACTGATGAAAGATTGTATTGCTGCAATGTTTATT
GATCCAGTGCCACTCTGAACTCA
INFORMATION FOR NO: lOB (SOSAyWS)
SEQliENCE CHARACTERISTICS:
LENGTH: 254
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY OLEFIN SYNTHASE.
SEQliENCE DISCRIPTION FOR NO: lOB

MPVHATPAAESQIISMPEWRRTANFKPSVWGDRFANYAEDIITQTQMQEQVEEL
KQVRKEVFTNAADDSSHQLKPIDEIQRLGVAYHFESEIDQALERIHETYQDIHDGG
DLYNVALRFRLLRRHGYNVSCDVFNKFIODTNGDYKIiSLVTDLSGMLSFYEAAHL
RVHGEKLLEEALVFTTTHLQSASAKSSLLKTQITEAVERLLKTMERLGARRYMSIY
QDEASYSENLLKLAia.DFNV\^QCLHKKELSDIP
INFORMATION FOR NO: IOC (SOSAAVS)
SEQliENCE CHARACTERISTICS:
LENGTH: 554
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY OLEFIN SYNTHASE.
Deleting the CC insertion
SEQliENCE DISCRIPTION FOR NO: IOC
MPVHATPAAESQliSMPEWRRTANFKPSVWGDRFANYAEDIITQTQMQEQVEEL
KQVRI^EVFTNAADDSSHQLKPIDEIQRLGVAYHFESEIDQALERIHETYQDIHDGG
DLYNVALRFRLLRRHGYNVSCDVFNKFKDTNGDYKKSLVTDLSGMLSFYEAAHL
RVHGEKli^EALVFirrHLQSASAKSSLliCrQITEAVERLLimVIERLGARRYMSIY
QDEASYSENliJO^AKLDFNVVQCLffl-aaELSDILRWYKELDFARRMPFARDRIVEL
FFWLAGIYFEPEYVFRRfflLTKLTEnrVMDDMYDAFGTFEELVNLTEAIDRWDASC
MDQLPDYMQPFYITLLDVIDEVEEELTKQGRSYRIHYAKEEVIKNQARLYFAEARW
FHEGCTPKMDEYMRVAASSVGNTMLSWSLVGMGDIITKFEFEWLTNEPiaLRAS
NTIFRLMDDIAGYKFEKERGHVASSIDCYMNEYGVSEQETIDIFNKRIVDSWKDIN
EEFLRPTAAPWVIJ^VLNLTRVVDLLYiaiGDAFrHVGiaJVIKDCL^yVMFIDPVPL

INFORMATION FOR NO: llA (SOSAMA)
SEQliENCE CHARACTERISTICS:
LENGTH: 2605
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY OLEFIN SYNTHASE.
INCLliDES A STOP CODON.
SEQliENCE DISCRIPTION FOR NO: llA
CTAATACGACTCACTATAGGGCAAGCAGTGGTAACAACGCAGAGTACGCGGGGA CATTTGATTCATAGTTATTAGATTGTGTTTTTCCGTCCAGTTAGGTTTAAGGATTA TACACTCGTTTAATGTATTGTTAGAACGGTGATTGTGTGCTTAGTTAATAGATTTT GCTTTTATTCAAGAGCGTAGGGTTCAATTTGAGTATGCATGTTCTTTTATCTTTAG CTTTTATTATGGAATTTTTATAAAATGTTATAATATTAATTTCTTAATGAGTAGTTA AATTACGTGATTATTTGATTTTTTTAATCTAAAATGTGATATGTAAAATATAGAAG AAAAAAAATTTAAAAACTTTCAGAAATTTTTTAAATTCTTTTAG CCCACCCAAACC TAAAATCCTAGGTCCGCCGTCGATGCAAAGTACAAATAGAAACATGTCTTTCTCA GTCATGAATCATGTCATCATGATATTGATAGATGATGTCGTTTAGCAATAAAGGG CTGTTCTGCGGTTAAAATATAAACATCTTCCGATCTTATTATTTACAACAACAAAA AATCTTCCAAACTCAATTATCAGCATCTGTATCAGATCTGCATGGAGTCCCCTATA AATATATGATCATAGCAGCAATATACTTCATACTTGAAGAAAAAGCTATAGCTAG TCCACAAGTGCAGAAAGTTAATCATGCCTGTCCATGCTACTCCAGCAGCTGAATC CCAGATCATCTCTATGCCGGAAGTTGTTCGGCGCACAGCAAATTTTAAACCTAGC GTTTGGGGAGATCGGTTTGCTAACTATGCCGAAGACATTATAACTCAAACTCAAA TGCAAGAACAAGTTGAGGAGCTGAAACAAGTAGTGAGGAAGGAAGTATTCACTA ATGCTGCTGATGATTCTTCACATCAACTGAAGCTAATTGATGAAATCCAGCGCCT CGGTGTGGCTTACCATTTCGAAAGCGAAATAGATCAAGCCCTGGAACGTATACAT GAGACATATCAAGATATTCATGATGGTGGTGATCTGTACAATGTTGCTCTTCGTT I TTCGGCTACTCAGGCGACATGGATATAATGTTTCCTGCGATGTATTCAACAAGTT CAAAGATACTAATGGTGACTACAAGAAAAGCTTGGTCACTGATCTTTCTGGTATG CTGAGCTTTTATGAGGCGGCCCATCTGAGGGTGCATGGAGAAAAATTACTTGAAG

AGGCTCTGGTTTTTACCACCACTCATCTCCAGTCAGGAAGTGCAAAAAGCTCTTT
GCTGAAAACACAAATAACTGAAGCCGTAGAGAGACTACTAAAAACTATGGAGAG
GTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCATACAGTGAA
AATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACAAAAA
GGAACTCAGTGACATTCCCTAAGATGGTACA.\GGAACTGGACTTTGCAAGGAGG
ATGCCTTTTGCTCGAGATAGGATCGTGGAGTTGTTCTTTTGGATAGCAGGAATAT
ATTTCGAACCTGAATACGTCTTTAGGAGACACATTCTGACTAAACTGATTGAGAT
AACAACAGTAATGGATGATATGTATGATGCATTCGGTACATTCGAAGAACTCGTC
AACTTGACTGAAGCAATTGACAGGTGGGATGCAAGTTGCATGGATCAACTGCCA
GACTATATGCAACCATTTTATATTACACTTCTGGATGTTATCGATGAAGTTGAAGA
GGAGCTGACAAAGCAAGGAAGATCTTACCGAATTCACTACGCAAAAGAAATTATG
AAGAATCAAGCCAGGCTCTACTTCGCTGAGGCCAGATGGTTCCACGAAGGATGC
ACCCCAAAAATGGATGAGTATATGCGAGTTGCGGCATCTTCTGTCGGTAACACCA
TGCTTTCCGTCGTGTCTTTAGTAGGCATGGGAGACATTATAACAAAATTTGAATT
CGAGTGGCTGACCAATGAGCCTAAAATCCTTAGAGCTTCGAATACCATATTTAGG
CTTATGGATGACATTGCTGGGTACAAGTTTGAGAAAGAGAGAGGGCATGTTGCTT
CAAGTATTGATTGCTACATGAATGAATACGGGGTTTCAGAGCAAGAGACAATTGA
TATCTTCAACAAACGAATTGTGGATTCGTGGAAGGATATAAACGAAGAGTTTCTG
AGACCCACTGCTGCTCCAGTCCCTGTGCTTAATCGTGTTCTTAACCTAACCCGAG
TGGTrGATCTGCTTTACAAAAGGGGAGATGCCTTCACGCATGTCGGAAAACTGAT
GAAAGATTGTATTGCTGCAATGTTTATTGATCCAGTGCCACTCTGAACTCATCGG
ATCAGTCATCACATTCAGTCTCCTGATGCTAGCGTTTGCTTTTTATTTGAATGTAT
TCTTGAATAAGACGATGCACCTCGATCAATTTGTGCTTCAGTGTTTCACGTACTG
ATGAGTCCTATCCTTTCTAGAAGAGGAACATCAATGTTGGTTTGCTAATAAAGCT
TTATTGTTTGAATGTCGGGTTGATAATTCTTAACTAATTATGTTGTCTAAAAAAAA AAAAAAAAAAAAA

INFORMATION FOR NO: IIB (SOSAMA)
SEQliENCE CHARACTERISTICS:
LENGTH: 255
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY OLEFIN SYNTHASE.
SEQliENCE DISCRIPTION FOR NO: IIB
MPVHATPAAESQIISMPEWRRTANFIvPSVWGDRFANYAEDIITQTQMQEQVEEL
KQWRIOIVFTNAADDSSHQLKLIDEIQRLGVAYHFESEIDQALERIHETYQDIHDG
GDLYNVALRFRLLRRHGYNVSCDVFNKFKDTNGDYKKSLVTDLSGMLSFiTEAAH
LRVHGEKLLEEALVFTTTHLQSASAKSSLLKTQITEAVERLLKTMERLGARRYMSI
YQDEASYSENLLKLAKLDFNWQCLHKIvEI^DIP
IN'FORMATION FOR NO: llC (SOSA/MA)
SEQliENCE CHARACTERISTICS:
LENGTH: 555
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY OLEFIN SYNTHASE.
Deleting the CC insertion.
SEQliENCE DISCRIPTION FOR NO: IIC
MPVHATPAAESQnSMPEVVRRTANFia>SVWGDRFANYAEDIITQTQMQEQVEEL
KQWRKEVFTNAADDSSHQLKLIDEIQRLGVAYHFESEIDQALERIHETYQDIHDG
GDLYNVALRFRLLRRHGYNVSCDVFNKFKDTNGDYKKSLVTDLSGMLSFYEAAH
LRVHGEia^LEEALVFTTTHLQSASAKSSLLKTQITEAVERLLKTMERLGARRYMSI
YQDEASYSENliJCLAKLDFNVVQCLHKKELSDILRWYKELDFARRMPFARDRIVE
IJ'FWIAGIYFEPEYVFRRHILTKIJEITTVMDDMYDAFGTFEELVNLTEAIDRWDAS
CMDQIJ»DYMQPFYITliJ)VIDEVEEELTKQGRSYRIHYAKEIMKNQARLYFAEAR
WFHEGCTPKMDEYMRVAASSVGNTMLSWSLVGMGDIITKFEFEWLTNEPKILRA

SNTIFRLMDDIAGYKFEKERGHVASSIDCYMNEYGVSEQETIDIFNKRIVDSWKDI NEEFLRPTAAPVPVLNRVLNLTRWDLLYKRGDAFTHVGiaMKDCIAAMFIDPVP
L
INFORMATION FOR NO: 12A (SOSV)
SEQliENCE CHARACTERISTICS:
LENGTH: 1973
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: WILD STRAWBERRY OLEFIN SYNTHASE
SEQliENCE DISCRIPTION FOR NO: 12A
ATGCCTGTCCATGCTACTCCAGCAGCTGAATCCCAGATCATCTCTAAGCCGGAAG
TTGTTCGGCGCACAGCAAATTTTAAACCTAGCGTTTGGGGAGATCGGTTTGCTAA
CTATGCCGAAGACATTATAACTCAAACTCAAATGCAAGAACAAGTTGAGGAGCTG
AAACAAGTAGTGAGGAAGGAAGTATTCACTAATGCTGCTGATGATTCTTCACATC
AACTGAAGCTAATTGATGAAATCCAGCGCCTCGGTGTGGCTTACCATTTCGAAAG
CGAAATAGATCAAGCCCTGGAACGTATACATGAGACATATCAAGATATTCATGAT
GGTGGTGATCTGTACAATGTTGCTCTTCGTTTTCGGCTACCTAGGCGACATGGAT
ATAATGTTTCCTGCGATGTATTCAACAAGTTCAAAGATACTAATGGTGACTACAA
GAAAAGCTTGGTCACTGATCTTTCTGGTATGCTGAGCnTTATGAGGCGGCCCAT
CTGAGGGTGCATGGAGAAAAATTACTTGAAGAGGCTCTGGTTTTTACCACCACTC
ATCTCCAGTCAGCAAGTGCAAAAAGCTCTTTGCTGAAAACACAAATAACTGAAGC
CGTAGAGAGACCTCTACTAAAAACTATGGAGAGGTTAGGTGCTCGGGGTTACATG
TCAATATATCAAGATGAAGCTTCATACAGTGAAAATTTACTGAAACTTGCAAAATT
AGATTTTAATGTTGTTCAGTGTTTACACAAAAAGGAACTCAGTGACATTCTAAGAT
GGTACAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCTCGAGATAGGATCG
TGGAGTTGTTCTTTTGGATAGCAGGAATATATTTCGAACCTGAATACGTCTTTGG
GAGACACATTCTGACTAAACTGATTGAGATAACAACAGTAATGGATGATATGTAT
GATGCATTCGGTACATTCGAAGAACTCGTCATCTTGACTGAAGCAATTGACAGGT
GGGATGCAAGTTGCATGGATCAACTGCCAGACTATATGCAACCATnTATATAAC

ACTTCTGGATGTTATCGATGAAGTTGAAGAGGAGCTGACAAAGCAAGGAAGATCT
TACCGAATTCACTACGCAAAAGAAATTATGAAGAATCAAGCCAGGCTCTACTTCG
CTGAGGCCATATGGTTCCACGAAGGATGCACCCCAAAAATGGATGGGTATATGC
GAGTTGCGGCATCTTCTGTCGGTAACACCATGCTTTCCGTCGTGTCTTTAGTAGG
CATGGGAGACATTATAACAAAATTTGAATTCGAGTGGCTGACCAATGAGCCTAAiV
ATCCTTAGAGCTTCGAATACCATATTTAGGCTTATGGATGACATTGCTGGGTACA
AGTTTGAGAAAGAGAGAGGGCATGTTGCTTCTAGTATTGATTGCTACATGAATGA
ATACGGGGTTTCAGAGCAAGAGACAATTGATATCTTCAACAAACGAATTGTGGAT
TCGTGGAAGGATATAAACGAAGAGTTTCTGAGACCCACTGCTGCTCCAGTCCCTG
TGCTTAATCGTGTTCTTAACCTAACCCGAGTGGn^GATCTGCTTTACAAAAGGGG
AGATGCCTTCACGCATGTCGGAAAACTGATGAAAGATTGTATTGCTGCAATGTTT
ATTGATCCAGTGCCACTCTGAACTCATCGGATCAGTCATCACATTCAGTCTCCTG
ATGCTAGCGTTTGCTTTTTATTTGAATGTATTCTTGAATAAGACGATGCACCTCGA
TCAATTTGTGCTTCAGTGTTTCACGTACTGATGAGTCCTATCCTTTCTAGAAGAG
GAACATCAATGTTGGTTTGCTAATAAAGCTTTATTGTTTGAATGTCGGGTTGATA
ATTCTTAACTAATTATGTTGTCTACTTTGTACTTl'CAAACTCAATCTCAATACAGA
ATTTATAGTGTACGAACTAAAAAAAAAAAAJ\AAAAAAAAAAAAAAA
INFORMATION FOR NO: I2B (SOS\') SEQliENCE CHARACTERISTICS: LENGTH: 556 TYPE: Peptide STRANDNESS: Single TOPOLOGY: Linear
OTHER INFORMATION: WILD STRAWBERRY OLEFIN SYNTHASE SEQLIENCE DISCRIPTION FOR NO: 12B
MPVHATPAAESQIISKPEWRRTANFKPSVWGDRFANYAEDIITQTQMQEQVEELK QWRKEVFTNAADDSSHQLKLIDEIQRLGVAYHFESEIDQALERIHETYQDIHDGG
DLYNVAIilFRII.RRHGYNVSCDVFNKFia)TNGDYiaCSLVTDI^GMI^FYEAAHL RVIIGEKLIJJEALVFTTrHLQSASAKSSLLierQITEAVERPLLKTMERLGARRYMSI

YQDEASYSENLLiaAKLDFNVVQCLHia^LSDILRWYKELDFAERMPFARDB.IVE
LFFVnAG]YFEPEYVrFGRHILTia.IEITTVMDDMYDAFGTFEELVILTEAIDRWDAS
CMDQLPDYMQPFYITLLDVIDEVEEELTKQGRSYRIHYAIffilMIvNQARLYFAEAIW
FHEGCTPKMDGYMRVAASSVGNTMLSWSLVGMGDIITKFEFEWLTNEPKILRAS
NTIFRLMDDIAGYKFEKERGHVASSIDCYMNEYGVSEQETIDIFNKRIVDSWKDIN
EEFLRPTAAPVPVLNRVLNLTRVVDLLYiaiGDAFTHVGI INFORMATION FOR NO: 13A (SOSV1AV76)
SEQliENCE CHARACTERISTICS:
LENGTH: 289
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 13A
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCATACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA
AAAAGGAACTCAGTGACATTCTAAGGTAAATTAAGCCATCGATCTTATAGTTAAT
TAGTATATACATATACAAGATAAGTTATAACCTAATATTGTTCTAAATATACTAGA
TGGTACAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCTCGAGATAGGATC GTGGAGTTGTTCT
INFORI^LVTION FOR NO: 13B (SOSV1/W76)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR N0:13B

AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCATACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAAGATGGTACAAGGAACTGGACTTTGCAAGGA GGATGCCTTTTGCTCGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 130 (SOSV1AV76)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of wild strawberry.
SEQliENCE DISCRIPTION FOR NO: 13C
RLGARRYMSrYQDEASYSENLLKIAKLDFNVVQCLHKKELSDILRWYICELDFARR MPFARDRIVELF
INFORMATION FOR NO: 14A (SOSV2AV93)
SEQliENCE CHARACTERISTICS:
LENGTH: 289
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 14A
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCATACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTrTACACA AAAAGGAACTCAGTGACATTCTAAGGTAAATTAAGCCATCGATCTTATAGTTAAT TAGTATATACATATACAAGATAAGTTATAACCTAATATTGTTCTAAATATACTAGA

TGGTACAAGGAACTGGACTTTGCAAGGAGGATGCCCTTTGCTCGAGATAGGATC GTGGAGTTGTTCT
INFORMATION FOR NO: 14B (SOSV2AV93)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 14B
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCATACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAAGATGGTACAAGGAACTGGACTTTGCAAGGA GGATGCCCTTTGCTCGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 14C (SOSV2AV93)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of wild strawberry.
SEQliENCE DISCRIPTION FOR NO: 14C
RLGARRYMSIYQDEASYSENLliOli^VKLDFNVVQCLmavELSDILRWYKELDFARR MPFARDRTVELF

INFORMATION FOR NO: 15A (SOSV3AV90)
SEQliENCE CHARACTERISTICS:
LENGTH: 300
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Inclliding an intron. SEQliENCE DISCRIPTION FOR NO: 15A
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA
AAAAGGAACTCAGTGACAll'CTAAGGTAAACTAAGCCATCGATCTTATAGCTATT
AGTTGTATGTATATGTATACAAGATAAGTAATAACCTTCTAATATTGCTCTATATA
CTATATATAGATGGTATAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCACG
AGATAGGATCGTGGAGTTGTTCr
INFORMATION FOR NO: 15 B (SOSV3AV90)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA~
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 15B
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAAGATGGTATAAGGAACTGGACTTTGCAAGGAG GATGCCTTTTGCACGAGATAGGATCGTGGAGTTGTTCT

INFORMATION FOR NO: 15C (SOSV3AV90)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of wild strawberry.
SEQliENCE DISCRIPTION FOR NO: 15C
RLGARRYMSIYQDEASHSENLLKLAKLDFNWQCLHKKELSDILRWYICELDFARR MPFARDRIVELF
INFORMATION FOR NO: 16A (SOSV4AV79)
SEQliENCE CHARACTERISTICS:
LENGTH: 289
TYPE:
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 16A
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCATACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA
AAAAGGAACTCAGTGACATTCTAAGGTAAATTAGGCCATCGATCTTATAGTTAAT
TAGTATATACATATACAAGATAAGTTATAACCTAATATTGTTCTAAATATACTAGA
TGGTACAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCTCGAGATAGGATC
GTGGAGTTGTTCT

INFORMATION FOR NO: 16B (SOSV4A^r79)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: I6B
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCATACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAAGATGGTACAAGGAACTGGACTrrGCAAGGA GGATGCCITTTGCTCGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 16C (SOSV4/W79)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of wild strawberry.
SEQliENCE DISCRIPTION FOR NO: 16C
RLGARRYMSIYQDEASYSENLLiaAiaj)FNWQCLHKKELSDILRWYKELDFARR MPFARDRIVELF

INFORMATION FOR NO: 17A (SOSV5/W84)
SEQliENCE CHARACTERISTICS:
LENGTH: 300
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Inclliding an iatroa.
SEQliENCE DISCRIPTION FOR NO: 17A
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA
AAAAGGAACTCAGTGACATTCTAAGGTAAACTAAGCCATCGATCTTATAGCTATT
AGTTGTATATATATGTATACAAGATAAGTAATAACCTTCTAATATTGCTCTATATA
CTATATATAGATGGTATAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCACG
AGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: I7B (SOSV5AV84)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of wild
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 17B
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAAGATGGTATAAGGAACTGGACTTTGCAAGGAG GATGCCTTTTGCACGAGATAGGATCGTGGAGTTGTTCT

INFORMATION FOR NO: 17C (SOSV5AV84)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of wild strawberry.
SEQliENCE DISCRIPTION FOR NO: 17C
RLGARRYMSIYQDEASHSENLLKLAKLDFNWQCLHKICELSDILRWYKELDFARR MPFARDRIVELF
INFORMATION FOR NO: ISA (SOSA1AV66)
SEQliENCE CHARACTERISTICS:
LENGTH: 291
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Inclliding an intron and a CC insertion.
SEQliENCE DISCRIPTION FOR NO: ISA
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCATACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTITACACA
AAAAGGAACTCAGTGACATTCCCTAAGGTAAATTAAGCCATCGATCTTATAGTTA
ATTAGTATATACATATACAAGATAAGTTATAACCTAATATTGTTCTAAATATACTA
GATGGTACAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCTCGAGATAGGA
TCGTGGAGTTGTTCT

INFORMATION FOR NO: 18B (SOSA1AV66)
SEQliENCE CHARACTERISTICS:
LENGTH: 206
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Not inclliding an intron. Inclliding a CC insertion.
SEQliENCE DISCRIPTION FOR NO: 18B
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCATACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA
AAAAGGAACTCAGTGACATTCCCTAAGATGGTACAAGGAACTGGACTTTGCAAGG AGGATGCCTTTTGCTCGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: ISC (SOSA1AV66)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNT:SS: single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of cliltivated strawberry. Deleted CC insertion
SEQliENCE DISCRIPTION FOR NO: 18C
RLGAERYMSIYQDEASYSENLLI^LAKLDFNWQCLHKIiELSDILRWYKELDFARR MPFARDRIVELF

INFORMATION FOR NO: 19A (SOSA2A?V68)
SEQliENCE CHARACTERISTICS:
LENGTH: 296
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 19A
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA
AAAAGGAACTCAGTGACATTCTAAGGTAAACTAAACAATCGATCTTATAGTTATT
AGTTGTGTATGTATACAAGATACGCAATAACCATCTAATATTGCTCTATATATGTA
CTATAGATGGTATAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCTCGAGAT
AGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 19B (SOSA2AV6S)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 19B
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGGTTCACACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAAGATGGTATAAGGAACTGGACTTTGCAAGGAG GATGCCTTTTGCTCGAGATAGGATCGTGGAGTTGTTCT

AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG
TGNAAAATTTACTCGAAACTTGCAy\AATTAGATTTTAATG'rTGTTCAGTGTTTACA
CAAAAAGGAACTCAGTGACATTCTAAGGTAAACT/\AACAATCGATCTTATAGTTA
TTAGTTGTGTATGTATACAAGATACGCAATAACCATCTAATATTGCTCTATATATG
TACTATAGATGGTATAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCCCGAG
ATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 20B (SOSA3AV46)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberrj'. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 20B
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAAGATGGTATAAGGAACTGGACTTTGCAAGGAG GATGCCTTTTGCCCGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 20C (SOSA3/W46)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of cliltivated strawberry.
SEQliENCE DISCRIPTION FOR NO: 20C

INFORMATION FOR NO: 19C (SOSA2/W68)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of cliltivated strawberry.
SEQliENCE DISCRIPTION FOR NO: 19C
RLGARRYMSIYQDEASHSENLLKLAKLDFNWQCLHKKELSDILRWYKELDFARR MPFARDRIVELF
INFORMATION FOR NO: 20A (SOSA3AV46)
SEQliENCE CHARACTERISTICS:
LENGTH: 298
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 20A

AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAGGATGGTATAAGGAACTGGACTTTGCAAGGA GGATGCCTTTTGCTCGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 21C (SOSA4AV59)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of cliltivated strawberry.
SEQliENCE DISCRIPTION FOR NO: 21C
RLGARRYMSIYQDEASHSENLLKLAKLDFNV^^QCLHKIvELSDILGW^TvELDFARR MPFARDRIVELF
INFORMATION FOR NO: 22A (SOSA5/W74)
SEQliENCE CHARACTERISTICS:
LENGTH: 296
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 22A
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAAGGTAAACTAAACAATCGATCTTATAGTTATT AGTTGTGTATGTATACAAGATACGCAATAGCCATCTAATATTGCTCTATATATGTA

RLGAKRYMSIYQDEASHSENLLKLAKLDFNWQCLHKKELSDILRWYKELDFARR MPFAKDRIVELF
INFORMATION FOR NO: 21A (SOSA4AV59)
SEQliENCE CHARACTERISTICS:
LENGTH: 296
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 21A
AGAGGrrAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA
AAAAGGAACTCAGTGACATTCTAGGGTAA^\CTAAACAATCGATCTTATAGTTATT
AGTTGTGTATGTATACAAGATACGCAATAACCATCTAATATTGCTCTATATATGTA
CTATAGATGGTATAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCTCGAGAT AGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 21B (SOSA4AV59)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 21B

RLGARRYMSIYQDEASHSENLLIOAIvLDFNWQCLHKKELSDILRWYIOlLDFARR MPFARDRIVELF
INFORMATION FOR NO: 23A (SOSA6/W56)
SEQliENCE CHARACTERISTICS:
LENGTH: 302
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 23A
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA
AAAAGGAACTCAGTGATATTCTAAGGTAAACTAAGCCATCGATCTTATAGCTATT
AGTTGTATATATATGTATACAAGATAAGTAATAACCnTTAATATTGCTCTATATA
TACTATATATAGATGGTATAAGGAACTGGACTTTGCAi\AGAGGATGCCTTTTGCT
CGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 23B (SOSA6/VV56)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 23B
GTTC
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA

CTATAGATGGTATAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCTCGAGAT AGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 22B (SOSA5AV74)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 22B
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA A\AAGGAACTCAGTGACATTCTAAGATGGTATAAGGAACTGGACTTTGCAAGGAG GATGCCTTTTGCTCGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 22C (SOSA5AV74)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of cliltivated strawberry.
SEQliENCE DISCRIPTION FOR NO: 22C

AAAAGGAACTCAGTGATATTCTAAGATGGTATAAGGAACTGGACTTTGCAAAGAG GATGCCTTTTGCTCGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 23C(SOSA6AV56)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of cliltivated strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 23C
RLGARRYMSIYQDEASHSENLLia.AI INFORMATION FOR NO: 24A (SOSA7AV61)
SEQliENCE CHARACTERISTICS:
LENGTH: 302
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER DfFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 24A
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG
TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA
AAAAGGAACTCAGTGACATTCTAAGGTAAACTAAGCCATCGATCTTATAGCTATT
AGTTGTATATATATGTATACAAGATAAGTAATAACCTTCTAATATTGCTCTATATA
TACTATATATAGATGGTATAAGGAACTGGACTTTGCAAGGAGGATGCCTTTTGCT CGAGATAGGATCGTOGAGTrGITCT

INFORMATION FOR NO: 24B (SOSA7/W61)
SEQliENCE CHARACTERISTICS:
LENGTH: 204
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA of cliltivated
strawberry. Not inclliding an intron.
SEQliENCE DISCRIPTION FOR NO: 24B
AGAGGTTAGGTGCTCGGCGTTACATGTCAATATATCAAGATGAAGCTTCACACAG TGAAAATTTACTGAAACTTGCAAAATTAGATTTTAATGTTGTTCAGTGTTTACACA AAAAGGAACTCAGTGACATTCTAAGATGGTATAAGGAACTGGACTTTGCAAGGAG GATGCCTTTTGCTCGAGATAGGATCGTGGAGTTGTTCT
INFORMATION FOR NO: 24C (SOSA7/W61)
SEQliENCE CHARACTERISTICS:
LENGTH: 67
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Translation of fragment from PCR on genomic
DNA of cliltivated strawberry.
SEQliENCE DISCRIPTION FOR NO: 24C
RLGARRYMSIYQDEASHSENLLIOAICLDFNVVQCLHKICELSDILRWYKELDFARR MPFARDRIVELF

INFORMATION FOR NO: 25A (H64TAR2)
SEQliENCE CHARACTERISTICS:
LENGTH: 1665
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOI/NEROLIDOL SYNTHASE WITH TAKGET SIGNAL
SEQliENCE DISCRIPTION FOR NO: 25A
TCTAACCTCATGCAGCTTACACATAAGAAGCAGCTGCCTACTTTTCAAAGACGGG
GCATTGCCGAAGATAGCTTGCTTCCCAGTTCTACTACTCCCATAAAGCCGATGAA
CGTTGAAACCAAGCATACTAGAACTATGGGTGACATTnTGTCCAACATTGTCAG
AAGTTGGAACTATTCAGAAATGTCTTAAGGAATGTAGCAGAGCTAGATGCCCTTG
AAGGTTTGAATATGATCGATGCTGTTCAAAGGCTAGGCATTGATTTCCACTTTCA
ACGAGAAATCGATGAAATTCTGCACAAGCAAATGAGTAATGTATCTGCCTCTGAT
GATCTTCATGAGGTTGCACTTCGCTTTCGACTACTGAGGCAACATGGTTACTTCG
TGCCTGAAGATGTGTTTAACAACTTCAAGGACAGCAAAGGAACGTTCAAGCAAGT
TCTGGGTGAAGACATCAAGGGATTGATGAGCTTATACGGAGCTTCGCAGCTAGG
TACAGAAGGAGAAGATACACTTGTTGAAGCTGAAAAGTTTAGTGGCCATCTGCTA
AAGACTTCTCTGTCACATCTTGATCATCATCATGCCAGAATTGTTGGCAATACATT
GAGGAATCCTCATCACAAAAGCTTGGCCTCATTCATGGCCAGGAACTTTTTCGTT
ACTTCTCAAGCCACCAATTCATGGTTAAATTTGCTAAAAGACGTAGCAAAAACAG
ATTTCAATATGGTCCGGTCTCTGCATCAGAATGAAATAGTTCAAATTTCCAAATG
GTGGAAGGAGCTTGGATTGGCTAAGGAACTGAAGTTTGCAAGAGATCAACCACA
GAAATGGTACATTTGGTCCATGGCATGCCTAACAGATCCAAAGTTATCAGAGGAG
AGGGTTGAGCTCACAAAACCCATTTCTTTTGTCTATTTGATAGATGACATTTTCGA
TGTTTATGGAACTCTTGATGACCTCATTCTCTTCACAGAAGCTGTTAATAGATGG
GAAATTACTGCTATAGACCACTTACCAGACTATATGAAGATATGCTTCAAGGCTC
TCTATGATATGACTAATGAAATCAGCTGCAAGGTCTATCAGAAGCATGGATGGAA
CCCCTTACAATCTTTGAAAATTTCGTGGGCGAGTCTTTGCAATGCATTTTTGGTG
GAAGCAAAATGGTTCGCATCTGGGCAGCTGCCGAAGTCAGAAGAGTACTTGAAG

AACGGCATCGTTTCTTCTGGGGTTAATGTGGTTCTAGTCCACATGTTTTTTATCTT
GGGTCAAAACATAACCAGAAAGAGTGTGGAGTTGTTGAATGAAACTCCAGCCAT
GATATCGTCCTCAGCAGCAATTCTTCGACTCTGGGACGATTTAGGCAGTGCAAAG
GATGAGAACCAGGATGGGAACGATGGGTCGTATGTAAGGTGCTACTTAGAGGAA
CATGAAGGCTGTTCCATTGAGGAGGCACGAGAAAAGACGATTAATATGATTTCAG
ATGAATGGAAGAAACTGAACAGAGA.\CTGCTCTCTCCAAATCCATTTCCAGCAAC
AATCACATTGGCTTCTCTTAATCTCGCAAGAATGATCCCCTTGATGTATAGCTACG
ATGGCAACCAATACCTTCCATCTCTTAAAGAGTATATGAAACTGATGTTGTATGA
GACTGTATCAATGTAA
INFORMATION FOR NO: 25B (H64TAR2)
SEQliENCE CHARACTERISTICS:
LENGTH: 554
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE WITH TARGET SIGNAL
SEQliENCE DISCRIPTION FOR NO: 25B
SNLMQLTHKKQLPTFQRRGIAEDSLLPSSTTPIKPMNVETKHTRTMGDIFVQHCQ
IvLELFRNVLRNVAELDALEGLNMIDAVQRLGIDFHFQREIDEILHKQMSNVSASD
DLHEVALRFRLLRQHGYFVPEDVFNNFKDSKGTFKQVLGEDIKGLMSLYGASQL
GTEGEDTLVEAEKFSGHLLKTSLSHLDHHHARIVGNTLRNPHHKSLASFMARNFF
VTSQATNSWLNLXJmVAKTDFNNIVRSLHQNEIVQISKWWKELGLAKELKFARDQ
PQKWYIWSMACLTDPIOSEERVELTKPISFVYLIDDIFDVYGTLDDLILFTEAVNR
WEITAIDHLPDYMKICFKALYDMTNEISCKVYQKHGWNPLQSLKISWASLCNAFL
VEAKWFASGQLPKSEEYLKNGIVSSGVNWLVHMFFILGQNITRKSVELLNETPA
MISSSAAILRLWDDLGSAKDENQDGNDGSYVRCYLEEHEGCSIEEAREKTINMISD
EWIOOJ^Eli^PNPFPATITLASLNLARMIPLlVr^SYDGNQYLPSLIffiYMKLMLYE
TVSM

INFORMATION FOR NO: 26A (H64TAR6)
SEQliENCE CHARACTERISTICS:
LENGTH: 1665
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE WITH TARGET SIGNAL
SEQliENCE DISCRIPTION FOR NO: 26A
TCTAACCTCATGCAGCTTACACAAAAGAAGCAGCTTCCTACTTTTCAAAGACGGG
GCATTGCCGAAGATAGCTTGCTTCCCAGTTCTACTACTCCCATAAAGCCGATGAA
CGTTGAAACCAAGCATACTAGAACTATGGGTGACATTTTTGTCCAACATTCTCAG
AAGTTGGAACTATTGAAAACTGTCTTGAGGAATGTAGCAGAGCTAGATGCCCTTG
AAGGTTTGAATATGATCGATGCTGTTCAAAGGCTAGGCATCGATTACAACTTTCA
ACGAGAAATCGACGAAATTCTGCACAAGCAAATGAGTATTGTGTCTGCCTGTGAT
GATCTTCATGAGGTTGCACTTCGCTTTCGACTACTGAGACAACATGGTTACTTCG
TGCCTGAAGATGTGTTTAACAACTTCAAGGACAGCAAAGGAATGTTCAAGCAAGT
TCTGGGTGAAGACATCAAGGGATTGATGAGCTTATACGAAGCTTCGCAGCTAGGT
ACAGAAGGAGAAGATACACTTGTTGAAGCTGAAAAGTTTAGCGGCCATCTGCTAA
AGACTTCTCTGTCACATCTTGATCATCATCGAGCCAGAATTGTTGCAAATACATTG
AGGAATCCTCATCACAAAAGCTTGGCCCCATTCATGGCCAGGAACTTTTTCGTTA
CTTCTCAAGGCACCAATrCATGGTTAAATTTGCTAAAAGAAGTAGCAAAAACAGA
TTTCAATATGGTCCGGTCTCTGCACCAGAATGAAATAGTTCAAATTTCGAAATGG
TGGAAGGAGCTTGGATrGGCTAAGGAACTGAAGTrTGCAAGAGATCAACCACTG
AAATGGTACATTTGGTCCATGGCATGCCTGACAGATCCAAAGTTATCAGAGGAGA
GGGTTGAGCTCACAAAACCCGTCTCTTTTGTCTATTTGATAGATGACATTTTCGAT
GTTTATGGAACCCTTGATGAACTCATTCTCTTCACAGAAGCTGTTAATAGATGGG
AAATTACTGCTATAGACCACTTACCAGACTACATGAAGATATGCTTCAAGGCTCT
CTACGATATGACTAATGAATTCAGCAGCAAGGTCTATCTGAAGCATGGATGGAAC
CCCTTACAATCTTTGAAAATTTCGTGGGCGAGTCTTTGCAATGCATTTTTGGTGG
AAGCAAAATGGTTCGCATCTGGGCAGCTGCCGAAGTCAGAAGAGTACTTGAAGA

ACGGCATCGTTTCTTCTGGGGTACATGTGGGTCTAGTCCACATGTTTTTTCTCTT
GGGTCAAAACATAACCACAAAGAGTGTGGAGTTGTTGAATGAAACTCCAGCCATC
ATATCGTCCTCAGCAGCAATTCTTCGACTCTGGGACGATTTAGGAAGTGCAAAGG
ATGAGAACCAGGATGGGAACGATGGGTCGTATATAAGGTGCTACTTAGAGGAAC
ATGAAGGCTGTTCCATCGAGGAGGCACGAGAAAAGACGATTAATATGATTTCAGA
TGAATGGAAGAAACTGAACAGAGAACTGCTCTCTCCAAATCCATTTTCAGCAACA
TTCACATTGGCTTCTCTTAATCTCGCTAGAATGATCCCCATGATGTATAGCTACGA
TGGCAACCGATGCCTTCCTGATCTTAAAGAGTATGTGAAACTGATGTTGTATGAG
ACTGTATCAATGTAA
INFORMATION FOR NO: 26B (H64TAR6)
SEQliENCE CHARACTERISTICS:
LENGTH: 554
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE WITH TARGET SIGNAL
SEQliENCE DISCRIFTION FOR NO: 26B
SNLMQLTQKKQOTTQRRGIAEDSIJ^SSTTPIKPMNVETKHTRTMGDIFVQHSQ
KLELLKTVLRNVAELDALEGLNIVIIDAVQRLGIDYNFQREIDEILHKQMSIVSACDD
LHEVALRFRLLRQHGYFVPEDVFNNFKDSKGMFKQVLGEDffiGLMSLYEASQLG
TEGEDTLVEAEKFSGHLLKTSLSHLDHHRARIVANTLRNPHHICSLAPFMARNFFV
TSQATNSWLNLliCEVAKTDFNMVRSLHQNEIVQISKWWKELGLAKELKFARDQP
LKWYIWSMACLTDPKLSEERVELTKPVSFVYLIDDIFDVYGTLDELILFTEAVNRW
EITAIDHLPDY]VIKICFKALYDMTNEFSSK\'YLKHGWNPLQSLKISWASLCNAFLV
EAKWFASGQLPKSEEYLKNGIVSSGVHVGLVHIVIFFLLGQNITTKSVELLNETPAM
ISSSAAILRLWDDLGSAKDENQDGNDGSYIRCYLEEHEGCSIEEAREKTINMISDE
WTaa2>mELI^PNPFSATFriJ^IJSfLARMIPMMYSYDGNRCLPDLICEYVKLMLYET
VSM

INFORMATION FOR 27A (H64TAR4)
SEQliENCE CHARACTERISTICS:
LENGTH: 1865 bases
TYPE: cDNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOLWEROLIDOL SYNTHASE WITH TARGET SIGNAL
SEQliENCE DISCRIPTION FOR : 27A
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC
CTTCTTTAAAGTATTCAATCCTGCTCCAAAAAGCATCCCACGTATTGGCCAGTCTA
ACCTCATGCAGCTTACACATAAGAAGCAGCTGCCTACTTTTCAAAGACGGGGCAT
TGCCGAAGATAGCTTGCTTCCCAGTTCTACTACTCCCATAAAGCCGATGAACGTT
GAAACCAAGCATACTAGAACTATGGGTGACATTTTTGTCCAACATTGTCAGAAGT
TGGAACTATTCAGAAATGTCTTAAGGAATGTAGCAGAGCTAGATGCCCTTGAAGG
nTGAATATGATCGATGCTGTTCAAAGGCTAGGCATTGATTTCCACTTTCAACGA
GAAATCGATGAAATTCTGCACAAGCAAATGAGTAATGTATCTGCCTCTGATGATC
TTCATGAGGTTGCACTTCGCTTTCGACTACTGAGACAACATGGTTACTTCGTGCC
TGAAGATGTGTTTAACAACTTCAAGGACAGCAAAGGAACGTTCAAGCAAGTTCTG
GGTGAAGACATCAAGGGATTGATGAGCTTATACGAAGCTTCGCAGCTAGGTACA
GAAGGAGAAGATACACTTGTTGAAGCTGAAAAGTTTAGTGGCCATCTGCTAAAGA
CTTCTCTGTCACATCTTGATCATCATCATGCCAGAATTGTTGGCAATACATTGAGG
AATCCTCATCACAAAAGCTTGGCCTCATTCATGGCAAGGAACTTTTTCGTTACTAC
TCAAGCCACCAATTCATGGTTAAATTTGCTAAAAGACGTAGCAAAAACAGATTTC
AATATGGTCCGGTCTCTGCATCAGAATGAAATAGTTCAAATTTCCAAATGGTGGA
AGGAGCTTGGACTGGCTAAGGAACTGAAGTTTGCAAGAGATCAACCACAGAAAT
GGTACATTTGGTCCATGGCATGCCTAACAGATCCAAAGTTATCAGAGGAGAGGGT
TGAGCTCACAAAACCCATTTCTTTTGTCTATTTGATAGATGACATTTTCGATGTTT
ATGGAACTCTTGATGACCTCATTCTCTTCACAGAAGCTGTTAATAGATGGGAAAT
TACTGCTATAGACCACTTACCAGACTATATGAAGATATGCTTCAAGGCTCTCTAT

GATATGACTAATGAAATCAGCTGCAAGGTCTATCAGAAGCATGGATGGAACCCCT
TACAATCTTTGAAAATTTCGTGGGCGAGTCTTTGCAATGCATTTTTGGTGGAAGC
AAAATGGTTCGCATCTGGGCAGCTGCCGAAGTCAAAAGAGTACTTGAAGAACGG
CATCGTTTCTTCTGGGGTTAATGTGGTTCTAGTCCACATGTTTTTTATCTTGGGTC
AAAACATAACCACAAAGAGTGTGGAGTTGTTGAATGAAACTCCAGCCATGATATC
GTCCTCAGCAGCAATTCTTCGACTCTGGGACGATTTAGGAAGTGCAAAGGATGAG
AACCAGGATGGGAACGATGGGTCGTATGTAAGGTGCTACTTAGAGGAACATGAA
GGCTGTTCCATTGAGGAGGCACGAGAAAAGACGATTAATATGATTTCAGATGAAT
GGAAGAAACTGAACAGAGAACTGCTCTCTCCAAATCCATTTCCAGCAACAATCAC
ATTGGCTTCTCTTAATCTCGCAAGAATGATCCCCTTGATGTATAGCTACGATGGC
AACCAATGCCTTCCATCTCTTAAAGAGTATATGAAACTGATGTTGTATGAGACTG
TATCAATGTAATAATAATGACACTACTGGAAGTGGAGTTGAACTTCAAAGGTGGT
CAAGAGAAACAAGAAGCCTAAGCTGTGTCAGTGAGCTGTGACTTGGTTG
INFORMATION FOR 27B (H64TAR4)
SEQliENCE CHARACTERISTICS:
LENGTH: 578 amino acids
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE WITH TARGET SIGNAL
SEQliENCE DISCRIPTION FOR : 27B
MASSSRAFFKVFNPAPKSIPRIGQSNLMQLTHKKQLPTFQRRGIAEDSLLPSSTTPI
KPMNVETIOrniTMGDIFVQHCQKLELFRNVLRNVAELDALEGLNMIDAVQRLGI
DFHFQREIDEILHKQIVISNVSASDDLHEVALRFRLLRQHGYFVPEDVFNNFKDSKG
TFKQVLGEDffiGLMSLYEASQLGTEGEDTLVEAEKFSGHLLKTSLSHLDHHHARI
VGNTIJINPHHKSLASFMARNFFVTTQATNSWLNLLKDVAICTDFNMVRSLHQNEI
VQISKWWKELGIAKELKFARDQPQKWYIWSMACLTDPKLSEERVELTKPISFVYL
roDIFDVYGTIJDDIJIJTEAVNRWErrAroHLPDYlVQaCFKALYDlVrrNEISCKVYQ

HGWNPLQSLKISWASLCNAFLVEAKWFASGQLPKSKEYLK^GIVSSGVm^v'LVH
MFFILGQNITTKSVELLNETPAMISSSAAILRLWDDLGSAKDENQDGNDGSYVRCY
LEEHEGCSIEEAREICTINMISDEWKKLNRELLSPNPFPATITLASLNLARMIPLMYS YDGNQCLPSLIOIYMICLMLYETVSM
INFORMATION FOR: 28A (H64NORL)
SEQliENCE CHARACTERISTICS:
LENGTH: 2277 bases
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: CliLTIVATED STRAWBERRY
LINALOOL/NEROLIDOL SYNTHASE GENOMIC DNA FRAGMENT
SEQliENCE DISCRIPTION FOR : 28A
CTCCCACAGCTTCTTAGTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGC
CTTCTTTAAAGTATTCAATCCTCAAATTGCCTGCTACTTTTGAGATAGCTTGCTTC
CCAGTTCTATTACTATAAAGCCGATGAACGTTGAAACCAAGCATACTAGAACTAT
GGTAAAATTCTCGGAGCTTTCTCCGAAGTACATTTCTACAAAAGGGTAGAGCTAG
CTACTAAACAATAGTTAATTGACTGTGCCTTGCTTGCAGGGTGACATTTTTGTCCA
ACATTCTCAGAAGTTGGAACTATTGAAAACTGTCTTGAGGAATGTAGCAGAGCTA
GATGCCCTTGAAGGTTTGAATATGATCGATGCTGTTCAAAGGCTAGGCATCGATT
ACAACTTTCAACGAGAAATCGACGAAATCCTGCACAAGCAAATGAGTATTGTGTC
TGCCCGTGATGATCTTCATGAGGTTGCACTTCGCTTTCGACTACTGAGACAACAT
GGTTACTTCGTGCCTGAAGGTAAGTTTAATCACACGTATTATTTTTCGTTCGCTAA
ACGATATGAAACTATTTCATTCATAAACAGTTGTAAAACTTGTGTAGTAATACATA
TTTCTACGTGTTTGTTACAGATGTGTTTAACAACTTCAAGGACAGCAAAGGAACG
TTCAAGCAAGTTCTGGGTGAAGACATCAAGGGATTGATGAGCTTATACGAAGCTT
CGCAGCTAGGTACAGAAGGAGAAGATATACTTGTTGAAGCTGAAAAGnTAGCG
GCCATCTGCTAAAGACTTCTCTGTCACATCTTGATCATCATCGAGTCAGAATTGTT
GCAAATACATTGAGGAATCCTCATCACAAAAGCTTGGCCCCATTCATGGCCAGGA

ACTTTTTCGTTACTTCTCAAGCCACCAATTCATGGTTAAATTTGCTAAAAGAAGTA
GCAAAAACAGATTTCAATATGGTCCGGTCTCTGCACCAGAATGAAATAGTTCAAA
TGTCCAAGTAAGTTTGACAATGACTTCACCAGTGTCAGGACATTGATACTTTAATT
CACACAGGAGATACTTAGTGTAATTATGTGTATTTTTGACATTGTAGATGGTGOA
AGGAGCTTGGATTGGCTAAGGAACTGAAGTTTGCAAGAGATCAACCACTGAAAT
GGTACATTTGGTCCATGGCATGCCTGACAGATCCAAAGTTATCAGAGGAGAGGG
TTGAGCTCACAAAACCCATCTCTTTTGTCTATTTGATAGATGACATTTTCGATGTT
TATGGAACCCTTGATGACCTCATTCTCTTCACAGAAGCTGTTAATCGGTATATATG
AATTATATGCGTCAGTGATGAAATATAATCAGACTTGTTACCAATTTATGATTGAT
CAACAACCTATTGCATACATACAGATGGGAAATTACTGCTATAGACCACTTACCA
GACTATAtGAAGATATGCTTCAAGGCTCTCTATGATATGACTAATGAATTCAGCAG
CAAGGTCTATCTGAAGCATGGATGGAACCCCTTACAATCTTTGAAAATTTCGGTA
CATAACTATATATACAAACTGTGACTAATCTATCACATTTAACTTGATTATCGTTA
AAATCGTGAGCTTGGATTACAAGGTTTACATTGAGACCATTCATTCTGTAACTTCT
GTTGCAGTGGGCGAGTCTTTGCA.'VTGCATTTTTGGTGGAAGCAAAAATGGTrCGC
CTCTGGGAAGCTGCCGAAGTCAGAAGAGTACTTGAAGAATGGCATCGTTTCTTCT
GGGGTAAATGTGGTTCTAGTCCACATGTTTTTTCTCTTGGTCAGAACAAACCAGA
AAGAGTGTGGAGTTGTTGAATGAAACTCCAGCCATTATATCGTCCTCAGCAGCAA
TTCTTCGACTCTGGGACGATTTAGGAAGTGCAAAGGATGAGAACCAGGATGGGA
ACGATGGGTCGTATGTAAGGTGCTACTTAGAGGAACATGAAGGCTGTTCCATTGA
GGAGGCACGAGAAAAGACGATTAATATGATTTCAGATGAATGGAAGAAACTGAA
CAGAGAACTGCTCTCTCCAAATCCATTTCCAGCATCATTCACATTGGCTTCTCTTA
ATCTCGCAAGAATGATCCCCTTGATGTATAGCTACGATGGCAACCAATGCCTTCC
ATCTCTTAAAGAGTATATGAAACTGATGTTGTATGAGACTGTATCAATGTAA'ITAA
TAATAAGACTACCGGAAGTGGAGTTGAACTTCAAAGGTGGGTGGTCAAGAGAAA
CAAGAAGCCTAAG

INFORMATION FOR 29 (El)
SEQliENCE CHARACTERISTICS:
LENGTH: 227 bases
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA from both
sides of the methionine residlies
SEQliENCE DISCRIPTION FOR : 29A
GGCCGCGGGAATTCTATTCGCTGATCATAGATCAGATGGCATTGTCTACTCGGG
CCTTCTTTAAAGTATTCAATCCCCAAATTACTCCAAACAGTATCTCACATATTGGC
CAGTCTAACCTCATGCAGCTTACACAAAAGAAGCAGCTTCCTACTTTTCAAAGAC
GGGGCATTGCCGAAGATAGCTTGCTTCCCAGTTCTACTACTCCCATAAAGCCGAT
GCACGTT
INFORMATION FOR: 29B (El)
SEQliENCE CHARACTERISTICS:
LENGTH: 64 amino acids
TYPE: Peptide
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA from both
sides of the methionine residlies
SEQliENCE DISCRIPTION FOR : 29B
MAI^TRAFFKVFNPQITPNSISHIGQSNLMQLTQKKQLPTFQRRGIAEDSLLPSSTT
PffiPMHV

INFORMATION FOR: 30A (E2)
SEQliENCE CHARACTERISTICS:
LENGTH: 201 bases
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA from both
sides of the methionine residlies
SEQliENCE DISCRIPTION FOR : 30A
CCGCGGGAATTCGATTTGCTGATCATAGATCAGATGGCATAGTCTTTTCGGTCCC TCTTTAAAGTATTCAATCAAATTGCTCCAAAAATTAACTCACATGTTGGCCACTCT AAGAAGCAGCTGCCTGCTACTTTTCAAAGATGGGGCGTTGCCGAAGATAGCTTGC TTTCCAGTTCTAGTACTATAAAGCTGATGCACGTT
INFORMATION FOR: 31A (E3)
SEQliENCE CHARACTERISTICS:
LENGTH: 141 bases
TYPE: Genomic DNA
STRANDNESS: Single
TOPOLOGY: Linear
OTHER INFORMATION: Fragment from PCR on genomic DNA from both
sides of the methionine residlies
SEQliENCE DISCRIPTION FOR: 31A
CCGCGGGAATTCGATTTGCTGATCATAGATCAGATGGCATCGTCTTCTCGGGCCT TCTTTAAAGTATTCAATCCTCAAATTGCCTGCTACTTTTGAGAAAGCTTGCTTCCC AGTTCTATTACTATAAAGCCGATGCACGTT





We Claims
1. An isolated or recombinant nucleic acid or functional fragment thereof encoding a proteinaceous molecule essentially capable of isoprenoid bio-active compound synthesis when provided with a suitable substrate under appropriate reaction conditions.
2. A nucleic acid or functional fragment thereof according to claim 1 wherein said nucleic acid encodes a proteinaceous molecule essentially capable of synthesizing at least a monoterpene alcohol Hnalool when contacted with geranyl diphosphate (GPP) and/or at least a sesquiterpene alcohol nerolidol when contacted with farnesyl diphosphate (FPP) under appropriate reaction conditions.

3. A nucleic acid according to claims 1 or claim 2 wherein said proteinaceous molecule comprises a terpene synthase/cyclase.
4. A nucleic acid according to claim 3 wherein said proteinaceous molecule comprises a sesquiterpenoid synthase/cyclase.
5. A nucleic acid according claim 4 wherein said proteinaceous molecule comprises a nerolidol synthase/cyclase protein or functional fragment thereof.
6. A nucleic acid according to claim 5 wherein said nerolidol synthase/cyclase comprises nerolidol synthase.
7. A nucleic acid according to anyone of claims 1-3 wherein said sesquiterpene alcohol nerolidol comprises trans-nerolodoL
8. A nucleic acid according to anyone of claims 1-6 wherein said monoterpene alcohol comprises linalool.
9. A nucleic acid according to anyone of claims 1-8 wherein said nucleic acid encodes a proteinaceous molecule comprising an amino acid sequence or functional fragment thereof that is at least 50% identical to H64MUT sequence as shown in figure 2 or functional fragment thereof.
10.A nucleic acid or functional fragment thereof according to claim 1 wherein said nucleic acid encodes a proteinaceous molecule essentially capable of the synthesis

of at least one monoterpenoid when contacted with geranyl diphosphate (GPP)
under appropriate reaction conditions. 11.A nucleic acid according to claim 10 wherein said monoterpenoid comprises a-
pinene, B-pinene, sabinene, B-myrcene, a-phellandrene, B-pheUandrene, a-
terpinolene, a-terpineol, y-terpinene and/or dihydromyzcenol. 12,A nucleic acid according to claim 10 or claim 11 wherein said proteinaceous
molecule comprises a monoterpenene synthase/cyclase. 13.A nucleic acid according to anyone of claims 10-12 wherein said nucleic acid
encodes a proteinaceous molecule comprising an amino acid sequence or functional
fragment thereof that is at least 50% identical to SOSV sequence as shown in
figure 7. 14. A nucleic acid according to anyone of claims 1-13 obtainable from a eukaryote,
preferably a plant. 15.A nucleic acid according to anyone of claims 1-13 obtainable from an invertebrate
animal. 16. A nucleic acid according to anyone of claims 1-15 provided with a nucleic acid
encoding a plastid targeting signal. 17.A nucleic acid according to anyone of claims 1-15 provided with a nucleic acid
encoding a mitochondrial targeting signal. 18.A nucleic acid according to anyone of claims 1-15 encoding a proteinaceous
molecule essentially capable of isoprenoid bio-active compound synthesis in the
cytosol in a ceE when provided with a suitable substrate under appropriate
reaction conditions. 19.A nucleic acid according to anyone of claims 1-15 encoding a encoding a
proteinaceous molecule essentially capable of isoprenoid bio-active compound
synthesis in a plastid in a cell when provided with a suitable substrate under
appropriate reaction conditions. 20.A nucleic acid according to anyone of claims 1-15 encoding a encoding a
proteinaceous molecule essentially capable of isoprenoid bio-active compound
synthesis in a mitochondrium in a cell when provided with a suitable substrate
under appropriate reaction conditions.

21. A proteinaceous molecule encoded by nucleic acid according to anyone of claims 1-
20. 22.A vector comprising a nucleic acid according to anyone of claims 1-20. 23.A host comprising a nucleic acid according to anyone of claims 1-20 or a vector
according to claim 21. 24.A host according to claim 23 wherein said vector according to claim 22 and the
host expresses a nerolidol synthase/cyclase protein or polypeptide. 25,A host according to claim 23 wherein said vector according to claim 22 and the
host cell expresses a monoterpenene synthase/cyclase protein or polypeptide. 26. A plant or propagating material derived thereof which comprises a nucleic acid
according to anyone of claims 1-20 or a vector according to claim 22. 27.A method for producing a flavor, fragrance and/or bio-control agent comprising a)
transforming or transfecting a suitable host with at least one nucleic acid encoding
a proteinaceous molecule according to anyone of claims 1-20 b) expressing said
nucleic acid in the presence of a suitable substrate c) optionally isolating the
formed product. 28.A method according to claim 27 wherein said host comprises a microorganism,
plant cell or plant. 29.A method for producing a flavor, fragrance and/or bio-control agent in a ceU-free
lysatc expression system comprising expressing at least one nucleic acid encoding
a proteinaceous molecule according to anyone of claims 1-20 in the presence of a
suitable substrate and optionally isolating the formed product. 30.A compound such as a flavor, fragrance and/or bio-control agent obtainable by a
method according to anyone of claims 22-29. 31.Use of a compound according to claim 30 as an anti-microbial agent. 32.Use of a compound according to claim 30 in the processed food industry as a food
additive to modify the taste of syrups, ice-creams, frozen desserts, yogurts,
confectionery and like products. 33.Use of a compound according to claim 30 as a flavoring agent for oral medications
and vitamins. 34.Use of a compound according to claim 30 for providing additional flavor/aroma in
beverages, including alcoholic beverages.

35.Use of a compound according to claim 30 for enhancing or reducing plant
flavor/aroma/fragrance/scent. 36.Use of a compound according to claim 30 for enhancing the flavor/aroma of natural
products and/or synthetic and/or artificial products. 37.Use of a compound according to claim 30 for the industrial synthesis of nature
identical flavor/aroma substances and/or artificial flavor/aroma substances. 38XJse of a compound according to claim 30 as a pest control agent. 59. Use of a compound according to claim 30 for the biological control of the
interaction between plants and insects and/or the interaction between plants and
micro-organisms. 40,Use of a compound according to claim 30 for providing flavor/aroma in cosmetics,
creams, sun-protectant products, hair conditioners, cleaning products, personal
care products and health care products. 41.Use of a compound according to claim 30 as a disinfectant additive. 42.Use of a nucleic acid or firagment thereof encoding a proteinaceous molecule
according to anyone of claims 1-20 as a molecular marker or diagnostic tool. 43.Use of a proteinaceous molecule according to claim 21 for the production of an
antagonist. 44.Use of a proteinaceous molecule according to claim 21 for the production of an
antagonist wherein said antagonist is an antibody or functional equivalent thereo£ 45.Use of a proteinaceous molecule according to claim 21 production of an antagonist
to inhibit the synathesis of a compound according to claim 30. 46.Use of a compound according to claim 30 for the preparation of a composition. 47.A composition comprising a flavor, fragrance and/or bio-control agent according to
claim 30. 48.A composition according to claim 47 which is a pharmaceutical. 49.A composition according to claim 47 which is a nutraceutical. SO.Use of a composition according to claim 47 for augmenting or enhancing the aroma
and/or taste of food or non food products and/or protection of food or non food
products against fungal contamination and/or pest infestation. Sl.Use of a composition according to daim 47 for the biological control of pests. 52.Use of a composition according to claim 47 for the protection of stored products.

53.Use of a composition according to anyone of claim 47-49 for the prevention or
treatment of disease. 54.A method of treatment of disease comprising administering a composition
according to anyone of claims 47-49 with a carrier to a suitable recipient.

55. An isolated or recombinant nucleic acid or functional fragment thereof, substantially as hereinabove described and illustrated with reference to the accompanying drawings.


Documents:

1419-chenp-2003-assignement.pdf

1419-chenp-2003-claims.pdf

1419-chenp-2003-correspondnece-others.pdf

1419-chenp-2003-correspondnece-po.pdf

1419-chenp-2003-description(complete).pdf

1419-chenp-2003-drawings.pdf

1419-chenp-2003-form 1.pdf

1419-chenp-2003-form 13.pdf

1419-chenp-2003-form 26.pdf

1419-chenp-2003-form 3.pdf

1419-chenp-2003-form 5.pdf

1419-chenp-2003-pct.pdf

1419-chenp-2008 abstract duplicate.pdf

1419-chenp-2008 claims duplicate.pdf

1419-chenp-2008 description (complete) duplicate 2.pdf

1419-chenp-2008 description (complete) duplicate.pdf

1419-chenp-2008 drawings duplicate.pdf


Patent Number 223358
Indian Patent Application Number 1419/CHENP/2003
PG Journal Number 47/2008
Publication Date 21-Nov-2008
Grant Date 09-Sep-2008
Date of Filing 10-Sep-2003
Name of Patentee DE RUITER SEEDS R&D B V
Applicant Address LEEUWENHOEKWEG 52, 2661 CZ BERGSCHENHOCK
Inventors:
# Inventor's Name Inventor's Address
1 BOUWMEESTER, HENDRIK JAN KLOOSTERKAMPWEG 14, 6871 ZZ RENKUM,
2 AHARONI, ASAPH PINKAS 52 STREET, TEL-AVIV
3 JONGSMA, MAARTEN ANTHONIE HUSZARLAAN 46, 6708 MS WAGENINGEN,
4 VERHOEVEN, HENRICUS ADRIANUS MGR VAN DE VENSTRAAT 48, 5482 EN SCHIHNDEL,
PCT International Classification Number C12N 9/88
PCT International Application Number PCT/NL02/00089
PCT International Filing date 2002-02-12
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 01200488.3 2001-02-12 EUROPEAN UNION