RU2780626C2 - Pesticide protein toxins active against lepidoptera - Google Patents

Abstract

FIELD: biochemistry.

SUBSTANCE: invention relates to the field of biochemistry, in particular to a polynucleotide structure for giving resistance to an insect of Lepidoptera order. Protein toxic for black cutworm of Lepidoptera order; a vector expressing the specified polynucleotide structure; a host cell containing the specified polynucleotide structure; a transgenic plant containing the specified polynucleotide structure; a seed containing the specified polynucleotide structure; a biological sample containing the specified polynucleotide structure; and a composition containing the specified protein are also disclosed. A method for the production of seeds, using the specified vector, is disclosed.

EFFECT: invention allows for effective control of an insect pest of Lepidoptera order.

16 cl, 2 dwg, 5 ex

Description

LINK TO RELATED APPLICATION

[001] This application claims priority over U.S. Provisional Application No. 62/445313, filed January 12, 2017, which is incorporated herein by reference in its entirety.

INCLUDING A SEQUENCE LIST

[002] A file named "MONS434WO-sequence listing.txt" containing the Sequence Listing in a machine-readable format was created on January 11, 2018. This file is 166134 bytes in size as measured by MS-Windows® and is incorporated herein by reference in its entirety. This Sequence Listing consists of SEQ ID NO:1-SEQ ID NO:12.

FIELD OF TECHNOLOGY

[003] The invention relates generally to the field of proteins exhibiting inhibitory activity against insects, in particular to proteins exhibiting inhibitory activity against insect pests of agriculture from the order Lepidoptera (Lepidoptera), such as upsilon cutworm ("BCW ", Agrotis ipsilon).

BACKGROUND OF THE INVENTION

[004] Insect-inhibiting proteins produced by the bacterial species Bacillus thuringiensis (Bt) are known in the art. Certain Bt proteins can be used to control crop pests by spraying agriculturally acceptable formulations containing one or more of such proteins onto plants, by coating seeds with a composition formulated to contain an insecticidally effective amount of such proteins, or by effectively expressing said one or more proteins in plants/seeds.

[005] Only a few Bt proteins have been developed for use as transgenic traits for commercial use by farmers to control insect pests. Farmers rely on these proteins to get their prescribed spectrum of pest control and can continue to rely on broad spectrum chemicals when applied to leaves and soil for pest control. Some Lepidoptera insects such as Agrotis spp. and Striacosta spp. have proven to be particularly difficult to control using the transgenic insecticidal traits currently in use, including Cry1Ab, Cry1Ac, Cry1Fa, Cry2Ab, Cry2Ae, VIP3Aa and various other Bt toxins that are being used. less often. Therefore, there is a need for insect inhibitory proteins that are active against a wider range of insect pest species and for the use of toxins to overcome the development of pest resistance to existing pesticides, including the toxins currently used in pest control systems.

[006] This application describes a new family of proteins, variants and chimeric toxic protein constructs, each of which exhibits surprisingly effective insecticidal activity against Lepidoptera, especially against pests of Agrotis species, such as upsilon armyworm.

SUMMARY OF THE INVENTION

[007] A new group of insect-inhibiting polypeptides (the toxic proteins BCW 001, BCW 002 and BCW 003 and their pesticidal fragments) have been shown to have inhibitory activity against several lepidoptera pests of agricultural plants, in particular against the epsilon cutworm ( species Agrotis). Each protein can be used alone or in combination with each other and with other Bt proteins and insect-inhibiting agents in formulations and in planta, thereby offering alternatives to Bt proteins and chemical insecticides currently used in agricultural systems.

[008] According to the present invention, polynucleotide constructs are provided that comprise, in operable linkage, a heterologous promoter segment linked to a nucleotide sequence encoding an insecticidal protein having Cry1A characteristics that is smaller than a full-length Cry1A class toxic protein and that has an amino acid sequence from about position 1 to position 607 as indicated in SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO 6, SEQ ID NO: 8, SEQ ID NO: 10 and SEQ ID NO: 12, or an insecticidally active fragment thereof. Less than a full-length polypeptide that exhibits such insecticidal activity must exhibit an identity of at least about 100%, 99%, 98%, 97%, 96%, 95%, 94%, 94%, 92%, 91%, or 90% with the amino acid sequence BCW 001 as specified in SEQ ID NO: 2, from about position 1 to about position 606, or from about position 5 to about position 600. If full-length or significantly larger fragments of the toxin are to be used, the percentage of identity must be less restrictive, and represent a percentage identity from about 100 to about 95 to about 90 to about 85 or even up to 80% identity with respect to the sequences of full-length protein toxins, as indicated in SEQ ID NOS: 2, 4 and 6 because these toxic proteins exhibit commercially useful levels of biological activity when tested against epsilon cutworm larvae in food bioassays and when tested in planta on transgenic corn, cotton, and soybeans expressing such proteins.

[009] The invention also provides proteins that are toxic to the epsilon cutworm species, including proteins having the amino acid sequence as set forth in SEQ ID NO: 2 from position 256 to 606 (BCW 001 protein), and proteins having the amino acid sequence as indicated in any of SEQ ID NO: 4 and SEQ ID NO: 6 from amino acid position 257 to 607 (referred to herein as BCW 002 toxic protein and BCW 003 toxic protein, respectively).

[0010] Such insecticidal proteins also exhibit activity against Lepidoptera species selected from the group consisting of: Spodoptera frugiperda, Spodoptera exigua, Spodoptera litura, Mamestra configurata, Striacosta albicosta, Trichoplusia ni, Pseudoplusia includens, Anticarsia gemmatalis, Hypena scabra, Heliothis virescens, Agrotis subterranea, Pseudaletia unipuncta, Agrotis ipsilon, Agrotis orthogonia, Ostrinia nubilalis, Amyelois transitella, Crambus caliginosellus, Herpetogramma licarsisalis, Homoeosoma electellum, Elasmopalpus lignosellus, Cydia pomonella, Endopiza viteana, Grapholita molesta, Suleima helianthana, Plutella xylostella, Pectinophora gossypiella, Lymantria dispar , Alabama argillacea, Archips argyrospila, Archips rosana, Chilo suppressalis, Cnaphalocrocis medinalis, Crambus caliginosellus, Crambus teterrellus, Diatraea grandiosella, Diatraea saccharalis, Earias insulana, Egrias vittella, Helicoverpa armigera, Helicoverpa zea, Heliothisogram virescens, Herpetogram alis, Lobesia botrana, Pectinophora gossypiella, Phyllocnistis citrella, Pieris brassicae, Pieris rapae, Plutella xylostella, and Tuta absoluta.

[0011] The proteins of this invention may also exhibit biological activity against Lepidoptera species selected from the group consisting of: Spodoptera frugiperda, Spodoptera exigua, Spodoptera litura, Mamestra configurata, Striacosta albicosta, Trichoplusia ni, Pseudoplusia includens, Anticarsia gemmatalis, Hypena scabra , Heliothis virescens, Agrotis subterranea, Pseudaletia unipuncta, Agrotis ipsilon, Agrotis orthogonia, Ostrinia nubilalis, Amyelois transitella, Crambus caliginosellus, Herpetogramma licarsisalis, Homoeosoma electellum, Elasmopalectpus lignosellus, Cydia pomonella, Endopiza viteana, Grapholymaphorita molesta, Pelinosta helianth gossypiella, Lymantria dispar, Alabama argillacea, Archips argyrospila, Archips rosana, Chilo suppressalis, Cnaphalocrocis medinalis, Crambus caliginosellus, Crambus teterrellus, Diatraea grandiosella, Diatraea saccharalis, Earias insulana, Egrias vitella, Helicoverpa armigera, Helicoverpa zea virescens, Herpetogramma licarsisalis, Lobesia botrana, Pectinophora gossypiella, Phyllocnistis citrella, Pieris brassicae, Pieris rapae, Plutella xylostella, and Tuta absoluta.

[0012] The proteins of this invention and the constructs discussed herein can be incorporated into any vector, including plasmids, cosmids, phagemids, and the like.

[0013] Such vectors can be used to introduce the constructs of this invention into any number of host cells, including bacterial cells, yeast cells, and plant cells.

[0014] Host cells that are yeast cells can be Saccharomyces cereviseae or Saccharomyces pombe, etc. Bacterial host cells can be any number of known such host cells, including but not limited to E. coli, B thuringiensis, and other related bacilli. Plant host cells can be obtained from any number of plant species, including but not limited to plant cells from: alfalfa, banana, barley, beans, broccoli, cabbage, ornamental cabbage, carrots, cassava, castor beans, cauliflower, celery, chickpeas, Chinese cabbage, citrus, coconut, coffee, corn, clover, cotton, cucurbits, cucumber, Menzies, eggplant, eucalyptus, flax, garlic, grapes, hops, leeks, lettuce, frankincense pine, millet, melon, walnut, oat, olive tree, onion, ornamental plants, palm, pasture grasses, pea, peanut, pepper, pigeonpea, pine, potato, poplar, pumpkin, radish pine, radish, rapeseed, rice, rhizomes, rye, wild saffron , shrub, sorghum, southern pine, soybean, spinach, cucurbits, strawberry, sugar beet, sugar cane, sunflower, sugar corn, amber tree, sweet potato, switchgrass, tea, tobacco, tomato, triticale, sod grass, watermelon for and wheat.

[0015] Transgenic plants, in particular transgenic corn, cotton and soybean plants, can be obtained by introducing constructs according to this invention containing suitably modified polynucleotide sequences such as those indicated in SEQ ID NO: 7, SEQ ID NO: 9 and SEQ ID NO: 11, for example, into the genome of such plant cells and by reconstituting a fertile transgenic corn, soybean or cotton plant containing in its genome a genetic construct for expressing at least a protein toxin according to the invention, i.e. BCW 001, BCW 002 or a protein BCW 003 toxin. Such transgenic plants will have a polynucleotide construct introduced into their plant genome containing at least a heterologous promoter segment operably linked to a nucleotide sequence encoding a BCW 001, BCW 002 or BCW 003 insecticidal protein having an amino acid sequence selected from the group , consisting of SEQ ID N O: 8, SEQ ID NO: 10 and SEQ ID NO: 12, or their insecticidal protein fragment.

[0016] Seeds are also contemplated as a feature of the invention, wherein the seeds are obtained from such transgenic plants, and such seeds contain a detectable amount of a polynucleotide construct introduced into the plant's genome. Pollen, seeds, progeny plant cells, plant tissue and commercial products derived from each such transgenic plant will contain a detectable amount of the polynucleotide construct.

[0017] Any biological sample that contains at least a detectable amount of a polynucleotide construct encoding such a BCW 001, BCW 002, or BCW 003 protein will be within the scope of the invention.

[0018] Compositions are provided that provide an insecticidally effective amount of the BCW 001, 002, or 003 protein of the present invention and are intended to control Lepidoptera pest species. Such compositions may also contain an additional agent that is different from the toxic BCW protein. Such an agent would also be toxic to the same Lepidoptera species as the toxic BCW protein. The additional agent must be selected from the group of agents consisting of proteins or polypeptides whose amino acid sequence differs from the BCW protein, and may also be an RNA molecule that is toxic to the target insect pest (such as dsRNA, miRNA or small interfering RNA (siRNA)), and may also be an insecticidal chemical such as pyrethrin, an organophosphate pesticide, etc. Alternatively, the additional agent may be any compatible Cry or related toxic protein such as another Cry1A, Cry1Ab, Cry1Ac , Cry1A.105, Cry1Ae (but these are not preferred as they may not provide adequate stability control properties), or Cry1B, Cry1C, Cry1D, Cry1E, Cry1F, Cry1G, Cry1H, Cry1I, Cry1J, Cry1K, Cry1L, Cry2A, Cry2Ab, Cry2Ae, Cry4B, Cry6, Cry7, Cry8, Cry9, Cry15, Cry43A, Cry43B, ET35, ET66, TIC400, TIC800, TIC807, TIC834, TIC853, TIC1415, VIP3A, VI P3Ab, Axmi insecticidal proteins, DIG insecticidal proteins, eHIPs and VIP proteins, and any toxic protein known in the art to confer toxicity against upsilon cutworm larvae or applicable to other target Lepidoptera pests.

[0019] Such compositions may also contain additional pesticidal agents that are not necessarily toxic to the same target pest, such as additional agents selected from the group consisting of Cry1C, Cry3A, Cry3B, Cry34, Cry35, Cry51Aa1, ET29, ET33, ET34, ET70, TIC407, TIC417, TIC431, TIC901, TIC1201, TIC3131, 5307, DIG-10, Axmi184, Axmi205 and AxmiR1.

[0020] Also provided are methods for producing seeds that exploit the beneficial pesticidal properties of the proteins of the present invention. Such methods include a polynucleotide construct designed to express a BCW 001, BCW 002, or BCW 003 protein, or a protein showing at least about 90% identity with said protein, said method comprising planting one or more seeds that contain a polynucleotide expressing one or more of the BCW protein toxins of the invention, growing plants from such seeds, and then harvesting such seeds from the plants. The harvested seed will contain the polynucleotide construct and produce plants that are also resistant to epsilon cutworm infection.

[0021] Such plants can be corn, cotton, soybeans, or any other plants susceptible to Lepidoptera pest species that have been shown to be controlled by the proteins of this invention. Such plants are expected to be pre-produced transgenic plants that will benefit from the effects of the toxic properties of the proteins of this invention. Maize plants falling into this category include, but are not limited to, transformants selected from the group consisting of: , DAS59122-7, Bt10, Bt11, Bt176, CBH-351, DKB-83614-9, MIR162, MIR604, TC1507, TC6275, transformant 676, transformant 678, transformant 680, transformant 98140, DAS40278-9, DKB89790-5, MON21 -9, HCEM485, MON832, MON87427, NK603, T14, T25 and VCO01981-5. Soybean plants that fall into this category of transgenic plants are selected from the group consisting of: MON87751, DAS81419-2, MON87701, A2704-12, A2704-21, A5547-127, A5547-35, CV127, DAS44406-6, DAS68416- 4, DP356043, FG72, MON4032, ACS-GM003-1, MON87705, MON87708, MON89788, W62, W98 and GFM Cry1A. Transgenic cotton plants falling into this category are selected from the group consisting of: DAS24236-5, DAS21023-5, transformant 31707, transformant 31803, transformant 31807, transformant 31808, transformant 42317, BNLA-601, COT102, COT67B, transformant 1, GHB119, GK12, MON15985, MLS9124, MON1076, MON531, MON757, T303-3, T304-40, SGK321, transformant 19-51a, GHB614, LLCotton25, MON88701, MON88702, MON1445, MON1698 and MON88913. Transgenic sugarcane plants that fall into this category include the NXI-1T sugarcane transformant. Rice plants known in the art that would benefit from having a construct encoding such BCW protein toxins include rice transformants selected from the group consisting of: LLRICE06, LLRICE601, LLRICE62, GM-A17054 and GM-A17054.

[0022] Additionally, processed plant products are provided that contain a detectable amount of the disclosed recombinant polynucleotides. Such processed products include, but are not limited to: vegetable biomass, oil, meal, animal feed, flour, flakes, bran, fluff, husks, and processed seeds.

[0023] Methods for producing transgenic plants are also provided. Such methods include introducing a recombinant polynucleotide into a plant cell and selecting a transgenic plant that expresses an insect-inhibiting amount of the recombinant polypeptide encoded by the recombinant polynucleotide.

[0024] Other embodiments, features and advantages of the invention will be apparent from the following detailed description, examples and claims.

BRIEF DESCRIPTION OF GRAPHICS

[0025] In FIG. 1 shows a list of Lepidoptera pest species that have been tested in bioassays with the toxic proteins BCW 001, BCW 002, and BCW 003. "+" indicates mortality relative to buffer control; "-" indicates the absence of statistically significant observed mortality above the control level in the form of a buffer; "ND" means not tested with the corresponding toxic protein. BCW 001 has been shown to be lethal against Agrotis ipsilon (epsilon cutworm), Striacosta albicostsa, Helicoverpa zea, Ostrinia nubilalis (corn moth), Diatraea grandiosella, Trichoplusia ni, Pseudoplusia includens, and has not been shown to be lethal or growth retardant when tested against Spodoptera frugiperda (corn cutworm). Diatraea saccharalis has not been tested with BCW 001. BCW 002 has been shown to be lethal against Agrotis ipsilon (epsilon cutworm), Striacosta albicostsa, Helicoverpa zea (cotton cutworm), and Ostrinia nubilalis (corn moth), and has not been shown to be lethal or stunted when tested against Spodoptera frugiperda (corn armyworm). Diatraea saccharalis, Diatraea grandiosella (Southwestern corn moth), Trichoplusia ni (Cabbage cutworm), and Pseudoplusia includens (Soybean cutworm) were not tested with BCW 002. BCW 003 showed lethal effects against Agrotis ipsilon (Upsilon cutworm), Striacosta albicostsa , Helicoverpa zea (cotton bollworm), Ostrinia nubilalis (corn moth), Diatraea saccharalis, Diatraea grandiosella, Trichoplusia ni (cabbage bollworm), and Pseudoplusia includens (soy bollworm), and showed no lethal effects or growth retardation when tested against Spodoptera frugiperda (corn leaf owl).

[0026] In FIG. 2 shows the amino acid sequence alignment of BCW 001 (SEQ ID NO: 2, upper) versus BCW 002 (SEQ ID NO: 4, middle), and versus BCW 003 (SEQ ID NO: 6, lower); the asterisks below the three sequences represent amino acid differences at the selected position in at least one of the three different sequences.

BRIEF DESCRIPTION OF SEQUENCES

[0027] SEQ ID NO: 1 is the nucleotide sequence of natural B. thuringiensis strain EG4384 encoding the Lepidoptera toxic protein BCW 001.

[0028] SEQ ID NO: 2 is the established amino acid sequence of BCW 001 from the open reading frame as indicated in SEQ ID NO: 1.

[0029] SEQ ID NO: 3 is an artificial sequence encoding the chimeric Lepidoptera toxic protein BCW 002.

[0030] SEQ ID NO: 4 is the established amino acid sequence of BCW 002 from the open reading frame as set forth in SEQ ID NO: 3, wherein such BCW 002 protein consists of Cry1Ac domain I operably linked to BCW 001 domains II and III (from amino acid position 258 to amino acid position 606, as indicated in SEQ ID NO: 2) and operably linked to the Cry1Ac protoxin domain from amino acid positions 608 to 1177, as indicated in SEQ ID NO: 4 and SEQ ID NO: 6. SEQ ID NO: 4 (BCW 002) differs from SEQ ID NO: 6 (BCW 003) only at position 259, BCW 002 contains isoleucine at this position, BCW 003 contains threonine as in BCW 001.

[0031] SEQ ID NO: 5 is an artificial sequence encoding chimeric Lepidoptera toxic protein BCW 003.

[0032] SEQ ID NO: 6 is the established amino acid sequence of BCW 003 from the open reading frame as set forth in SEQ ID NO: 3, wherein such BCW 003 protein consists of Cry1Ac domain I operably linked to BCW 001 domains II and III (from amino acid position 258 to amino acid position 606, as indicated in SEQ ID NO: 2) and operably linked to the Cry1Ac protoxin domain from amino acid positions 608 to 1177, as indicated in SEQ ID NO: 4 and SEQ ID NO: 6. SEQ ID NO: 6 (BCW 003) differs from SEQ ID NO: 4 (BCW 002) only at position 259, BCW 002 contains isoleucine at this position, BCW 003 contains threonine, as in BCW 001.

[0033] SEQ ID NO: 7 is an artificial sequence encoding the BCW 001 protein for expression in plants.

[0034] SEQ ID NO: 8 is the established amino acid sequence of BCW 001 derived from SEQ ID NO: 7.

[0035] SEQ ID NO: 9 is an artificial sequence encoding the BCW 002 protein for expression in plants.

[0036] SEQ ID NO: 10 is the established amino acid sequence of BCW 002 derived from SEQ ID NO: 9.

[0037] SEQ ID NO: 11 is an artificial sequence encoding the BCW 003 protein for expression in plants.

[0038] SEQ ID NO: 12 is the established amino acid sequence of BCW 003 derived from SEQ ID NO: 11.

DETAILED DESCRIPTION OF THE INVENTION

[0039] An alternative to crop pest control by spraying formulations containing insecticidal proteins on plants/seeds is to insert polynucleotides encoding these proteins into the plant genome for expression in the plant or plant parts. Plants transformed with such polynucleotides have the insect resistance that these expressed proteins confer as transgenic traits.

[0040] In order to avoid developing or bypass insect resistance to currently used proteins, novel proteins with a different mode of action (DM) as well as broad spectrum and efficacy are needed to control Lepidoptera. One way to solve this problem is to sequence Bt genomes in the hope of discovering new insecticidal proteins. Another approach is to exchange segments from different Bt proteins to create new chimeric Bt proteins with insect inhibitory properties. The likelihood of randomly generating a chimeric protein with improved properties by recombining the domain structures of numerous natural insecticidal crystal proteins known in the art is extremely low (see, for example, A Strategy for Shuffling Numerous Bacillus thuringiensis Crystal Protein Domains; J. Economic Entomology, 97 (6 ): 1805-1813. 2004).

[0041] Disclosed herein are nucleotide sequences that encode insecticidal proteins, referred to herein as BCW proteins, that meet the need for an alternative DM, confer activity against a broader range of insect pests, and act to delay or prevent the development of resistance, especially when use in pest control upsilon cutworms (BCW).

[0042] BCW 001 was discovered as an open reading frame predicting an amino acid sequence having the characteristics of a Cry1A type protein after genome sequencing of Bacillus thuringiensis strain EG4384. The BCW 001 open reading frame (ORF) encoded a 1180 amino acid protein, and the protein was predicted to have many of the characteristics of Cry1 protein toxins, including an identifiable structure of domains I, II, and III, and a characteristic Cry1A-type protoxin domain on the carboxy-terminal half of the predicted protein. The predicted domain I amino acid sequence (residues 1 to about 258 of SEQ ID NO: 2) shows about 67% identity with domain I of the Cry1Ac protein toxin. The predicted amino acid sequence of domain II (residues from about 259 to about residue 459, as indicated in SEQ ID NO: 2) shows complete (100%) identity with domain II of Cry1Ai2. The predicted domain III amino acid sequence (residues from about 260 to about 606 as indicated in SEQ ID NO: 2) shows about 63% identity with the corresponding domain III residues in Cry1Ah2. The structure of the protoxin domain of the predicted BCW 001 protein (from about residue 607 to 1180 as indicated in SEQ ID NO: 2) shows about 96% identity with the corresponding residues in Cry1Aa9. Overall, this predicted full-length protein shows about 83% amino acid sequence identity with Cry1Ai, and the predicted toxin region from amino acid position 1 to about residue 607, as indicated in SEQ ID NO: 2, shows 76% identity with Cry1Ai1. It is difficult to assign this new toxic protein to a specific Cry1A class, and so the Bacillus thuringiensis nomenclature committee will be given the sequence and determine if this protein deserves its own separate and new class.

[0043] The BCW 001 protein was derived from a plasmid vector in a non-crystal-bearing strain of Bacillus thuringiensis, and spore crystal preparations were tested against a variety of Lepidoptera. See Fig. 1, column 2 for data. The facts indicate that this protein is not similar to any of the proteins with which it shares a source of origin, ie the toxic Cry1Aa, Cry1Ah or Cry1Ai proteins known in the art. None of the prior art proteins show any appreciable activity when tested against upsilon cutworms, however, this new protein BCW 001 was toxic in bioassays when tested against upsilon cutworms and unexpectedly showed toxic properties when tested against a group of other lepidoptera, such as set out in FIG. one.

[0044] In particular, in bioassays compared to an untreated insect control diet, BCW 001 protein was active against Striacosta albicosta ("WBC"), Helicoverpa zea ("CEW"), corn moth ("ECB", Ostrinia nubilalis), Diatraea grandiosella ("SWC"), Pseudoplusia includes ("SL"), Trichoplusia ni ("CLW"), and cutworms upsilon 1-3 larval stages ("BCW", Agrotis ipsilon).

[0045] As described further below, chimeric toxic proteins were generated using Cry1Ac domain I, and BCW 001 domains II and III (i.e., BCW 002 and BCW 003 as set forth in SEQ ID NO: 4 and SEQ ID NO: 6, respectively). ), and these chimeric toxic proteins have been introduced into corn and sugarcane plants. Both chimeric proteins were active in maize against BCW, WBC, CEW and SWC. BCW 003 was active against SCB in sugar cane.

[0046] The term "BCW protein" as used herein refers to any novel insect inhibitory protein that contains, consists of, is substantially homologous to, or is derived from any insect inhibitory polypeptide sequence: BCW 001 (SEQ ID NO: 2), BCW 002 (SEQ ID NO: 4) and BCW 003 (SEQ ID NO: 6) and their insect inhibitory segments, or combinations thereof, which provide activity against Lepidoptera, in particular, but not limited to, BCW , WBC and/or SCB. The polynucleotide encoding BCW 001 was derived from strain EG4384. The main toxic amino acid sequence for BCW 001 corresponds to amino acids from about position 28 to about position 606 to position 618, as indicated in SEQ ID NO: 2, and the main toxic amino acid sequence for BCW 002 and 003 corresponds to amino acids from about position 29 to about position 607 and up to about position 619, as indicated in SEQ ID NO: 4 and SEQ ID NO: 6, respectively.

[0047] In one embodiment, the proteins disclosed herein are related in primary structure to delta-endotoxin in length (about 1176-1180 amino acids), in protein length without protoxin (about 600 to about 619 amino acids), in length the main body of the toxin (about 591 amino acids) or by the presence of at least one BCW-specific segment.

[0048] Exemplary proteins were aligned to each other using the Clustal W algorithm, resulting in the number of amino acid matches for alignment pairs and percent amino acid identity for alignment pairs using these default parameters: Weight Matrix: Blosum; Penalty for opening a pass: 10.0; Pass renewal penalty: 0.05; Hydrophilic passes: On; Hydrophilic residues: GPSNDQERK; Skipping penalties for specific balances: Incl. The Clustal W algorithm is described in Thompson, J.D., Higgins, D.G. and Gibson, T.J. (1994) "CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, positions-specific gap penalties and weight matrix choice." Nucleic Acids Research, 22:4673-4680.

[0049] Other alignment algorithms are also available in the art and provide similar results to those obtained using Clustal W alignment.

[0050] The term "about" is used herein to describe that these segment boundaries may differ by 1, 2, 3, 4, 5, 10, 20, 25, 30, or 35 residues, depending on the sequence of the original proteins and their alignment with each other. To further describe the variability and configuration of the different segments, Tables 2 and 3 show the segment boundaries of BCW 002 and 003, and other chimeric proteins active against cutworm-upsilon.

[0051] The term "fragment" is used herein to describe continuous amino acid or nucleotide sequences that are shorter than the full amino acid or nucleotide sequence describing the toxic BCW protein.

[0052] The phrase "insect inhibitor" and "insecticide" are used interchangeably herein and refer to a protein, protein fragment, protein segment, or polynucleotide that results in any measurable inhibition of insect viability, growth, insect development, insect reproduction, insect feeding behavior , mating behavior of insects, and/or any measurable reduction in adverse effects caused by insect ingestion of a given protein, protein fragment, protein segment, or polynucleotide.

[0053] The terms "biological activity", "active", "activity", "effective", "efficacy" or variations thereof are used interchangeably herein to describe the effects of the proteins of this invention on target insect pests.

[0054] A crop is a self-propagating or cultivated plant whose product is harvested at some point in its growth stage. Non-limiting examples of such products are seed, pod, leaf, flower, stem, root, or any part thereof.

A biological sample derived from any plant tissue, bacterium, virus, or vector containing a polynucleotide or expressing a protein as illustrated herein, such as, but not limited to, a seed, boll, leaf, flower, stem, root, or any of them part, and containing a detectable amount of polynucleotide, protein, or both.

[0055] The phrase "detectable amount" is used herein to describe the minimum amount of a protein or polynucleotide disclosed herein that can be detected by standard analytical methods such as, but not limited to, polymerase chain reaction (PCR) and enzyme immunoassay ( ELISA), etc.

[0056] In one embodiment, the toxins described herein share a common function and exhibit insecticidal activity against Lepidoptera insect species.

[0057] BCW 001 segments confer activity against scoop-upsilon Cry1 chimeras that contain such segments. Examples of BCW 001 segments that confer activity against upsilon cutworms are shown in SEQ ID NO: 2 from about amino acid position 250 to about 606, and more specifically from about amino acid position 255 to about 606. Cry1 chimeras containing this amino acid segment corresponding to domains II and III of the toxic BCW 001 protein will often also have, in addition to the properties of the chimeric protein, toxic properties associated with the control of scoop-upsilon, and this has been tested in the frameworks Cry1A, Cry1B, Cry1C, Cry1D, Cry1E and Cry1F, in which the corresponding toxic construct had domain II and III components substituted with this region of amino acids from BCW 001, and in many cases anti-epsilon cutworm activity was unexpectedly retained in the chimeric construct (data not shown).

[0058] In one aspect of the invention, the pest controlled by the applicable toxic BCW protein is specifically an insect pest of the order Lepidoptera, including adults, pupae, larvae, and emerging larvae.

[0059] Insects of the order Lepidoptera (Lepidoptera) include, but are not limited to: insects of the Noctuidae family (caterpillars, cutworms, moths, heliothines), (e.g., corn fallowworm (Spodoptera frugiperda), (Spodoptera exigua), Mamestra configurata, Agrotis ipsilon cutworm, Trichoplusia ni, Pseudoplusia includens, Anticarsia gemmatalis, Hypena scabra, Heliothis virescens, Agrotis subterranea, Agrotis orthogonia, Pseudaletia unipuncta; Pyralidae), e.g. corn moth (Ostrinia nubilalis), Amyelois transitella, Crambus caliginosellus, Herpetogramma licarsisalis, Homoeosoma electellum, Elasmopalpus lignosellus; insects from the leafworm family (Tortricidae), e.g. apple codling moth (Cydia pomonella), Endopiza viteana, eastern codling moth (Grapholita molesta ), Suleima helianthana; and many other economically important Lepidoptera (e.g. , cabbage moth (Plutella xylostella), Pectinophora gossypiella, and gypsy moth (Lymantria dispar)). Other insect pests of the Lepidoptera order include, for example, Alabama argillacea, Archips argyrospila, Archips rosana and other Archips species, Chilo suppressalis (yellow rice moth), Cnaphalocrocis medinalis, Crambus caliginosellus, Crambus teterrellus, Diatraea grandiosella, Diatraea saccharalis , Earias insulana, Earias vittella, Helicoverpa armigera (cotton bollworm), Helicoverpa zea, Heliothis virescens, Herpetogramma licarsisalis, Lobesia botrana (cluster leafworm), Pectinophora gossypiella, Phyllocnistis citrella, Pieris brassicae (cabbage), Pieris rapae (turnip), Plutella xylostella (cabbage moth), Spodoptera exigua, Spodoptera litura, S. frugiperda, and Tuta absoluta (tomato miner moth).

[0060] The proteins disclosed herein can also be used to generate antibodies that specifically bind to BCW-specific toxic proteins and can be used to screen and search for other members of the BCW toxin family.

[0061] In one embodiment, exemplary polynucleotides that encode insect inhibitory proteins related to BCW 001 are shown in SEQ ID NOS: 1, 3, 5, 7, 9, and 11. Nucleotide sequences encoding these proteins can be used as probes and primers for screening to identify other members of the family using thermal or isothermal amplification and/or hybridization methods and other identification methods known to those skilled in the art.

[0062] According to an aspect of the invention, methods are provided for discovering related proteins, and such methods include sequencing Bt genomes, assembling sequencing data, identifying and cloning Bt genes encoding such pesticidal proteins, and expressing and testing novel Bt proteins to analyze pesticidal activity. . In another aspect of the invention, molecular methods are used to construct and clone commercially useful proteins, including chimeras of proteins from the pesticidal protein family, for example, chimeras can be assembled from segments of toxic BCW proteins to provide additional embodiments. The disclosed proteins can be aligned with each other and with other pesticidal Bt proteins, and segments of each such protein can be identified that are useful for substitution between the aligned proteins, resulting in the construction of chimeric proteins. Such chimeric proteins can be subjected to a bioassay with a pest and characterized for increased biological activity or broadening of the spectrum of target pests compared to the original proteins from which each such segment of the chimera was derived. The pesticidal activity of the polypeptides can be further engineered to be active against a particular pest or a broader range of pests by replacing domains or segments with those of other proteins.

[0063] In one embodiment, the proteins disclosed herein include functionally equivalent fragments (N- or C-terminal deletions) of the proteins disclosed herein.

[0064] BCW toxic proteins are provided herein. In some embodiments, BCW 001 related toxins can be isolated, formulated, in a transgenic microorganism or in a transgenic plant. , especially inhibitory activity against cutworms-upsilon and/or Diatraea saccharalis. Reference in this application to "isolated protein", or an equivalent term or phrase, is intended to mean that a protein is a molecule that is present alone or in combination with other compounds, but not in its natural environment. For example, toxic proteins according to the present invention and the like that are naturally found in the body are not considered "isolated" if they are in the body in which they are in a natural state. However, each will be "isolated" within the scope of this disclosure if the protein is not found in the organism in which it occurs naturally.

[0065] The term "operably linked", as used herein, refers to the connection of nucleic acid sequences or amino acid sequences such that one sequence can provide a desired function or compatible or useful property to the associated sequence.

[0066] Peptides, polypeptides, and proteins biologically functionally equivalent to BCW 001, BCW 002, and BCW 003 include, but are not limited to: amino acid sequences containing conservative amino acid substitutions in these protein sequences of the toxic BCW protein. In such amino acid sequences, one or more amino acids in the sequence are replaced by another amino acid(s), resulting in a silent or conservative substitution in the amino acid sequence.

[0067] Although the disclosed insect inhibitory polypeptides preferably comprise the protein sequence BCW 001, BCW 002, or BCW 003, fragments and variants of this sequence having the same or similar insect inhibitory activity as that of the insect inhibitory protein are also disclosed herein. document. For example, contiguous sequences of at least 30, 35, 38, 40, 45, 50, 55, 60, 65, 70, 75, 100, 150, 200, 500, 550 or more amino acids in a BCW-related inhibitory toxic protein activity against insects. In another embodiment, fragments of a BCW-related insect inhibitory toxic protein may contain amino acid substitutions, deletions, insertions, or additions to the BCW toxic protein sequence.

[0068] In one embodiment, the insect inhibitory polypeptide comprises an insect inhibitory segment from about residue 28 to about residue 618 of the BCW 001 protein sequence as set forth in SEQ ID NO: 2. Non-limiting examples include any of SEQ ID NO: 2, 4 or 6 or shorter fragments or variants having the same or similar insect inhibitory activity as that of that particular BCW 001 protein, either alone or operably linked to a chimeric protein. In another embodiment, segments having contiguous amino acid sequences of at least about 38 or more amino acids in any of SEQ ID NO: 2, 4, or 6 having insect inhibitory activity also yield a functional insecticidal protein. Insect-inhibiting toxic fragments of BCW 001 may also contain segments of at least 580 or 585 amino acid residues from a region of 591 amino acid residues corresponding to residues from about 28 to about 618 sequences, any of SEQ ID NO: 2, 4 or 6.

[0069] In some embodiments, fragments of the mature BCW 001 protein (mature refers to the 1180 amino acid protoxin form of the protein, cleaved by proteolysis in the gut of the insect pest to release only the N-terminal core of the toxin up to and including residue 607 near residue 618, releasing the active segment of a toxin containing, more or less, residues 1 to 606 or any number of residues from about 5 to about 618, as indicated in SEQ ID NO: 2, provided that the released segment exhibits toxic properties when exposed to cutworm larvae -upsilon) can be truncated forms in which one or more amino acids are removed from the N-terminus, C-terminus, midpoint of the protein, or combinations thereof, with insect suppression activity. These fragments may be natural or synthetic variants of BCW 001 and retain the insect inhibitory activity of the toxic BCW protein. In some embodiments, fragments of mature BCW 001, BCW 002, or BCW 003 proteins exhibit the pesticidal activity of the original protein molecules from which they are derived. A fragment or variant described herein may further comprise a domain designated herein that is responsible for the pesticidal activity of the protein. The truncated derivative having insect inhibitory activity is the toxic BCW protein corresponding to residues from about 28 to about 606 or up to about 618 of the sequence of the toxic BCW 001 protein as set forth in SEQ ID NO: 2, or to residues from about 29 to about 29 to about 607 and up to residue 619 of the toxic BCW protein as indicated in SEQ ID NO: 4 or SEQ ID NO: 6.

[0070] In yet another embodiment, truncated N-terminal deletion mutations include, but are not limited to, toxic BCW 001 proteins that lack amino acid residues from either the N-terminus and/or the C-terminus of the toxic protoxin-free portion, or the toxic core of toxic BCW proteins. For example, 1 to 6 N-terminal amino acid residues of the toxic core of the BCW 001 protein corresponding to residues 28 to 618 of SEQ ID NO: 2 or residues 29 to 619 of SEQ ID NO: 4 or 6 may be deleted. Truncated C-terminal deletion mutants of the toxic BCW protein corresponding to residues 28 to 618 of SEQ ID NO: 2 or residues 29 to 619 of SEQ ID NO: 4 or 6 include, but are not limited to, toxic BCW proteins in which missing from 1 to 6 C-terminal amino acid residues. In other embodiments, a toxic BCW protein with the corresponding residues 28-618 of SEQ ID NO: 2 or the corresponding residues 29-619 of SEQ ID NO: 4 or 6 may have both an N-terminal truncation of 1 to 6 amino-terminal residues, and a C -terminal truncation from 1 to 6 carboxyterminal residues.

[0071] In some embodiments, individual segments 1 to 6 of the CPR24719 protein, or a combination of segments 1 to 6, which confer anti cutworm-upsilon activity to a protein other than CPR24719-1, may also exhibit the same or a similar function.

[0072] Fragments and variants of the toxic BCW protein disclosed herein may have about 62% or more sequence identity, about 65%, 70%, 75%, 80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or greater sequence identity and about 99%, 99.5%, 100% amino acid sequence identity with the corresponding segments of the mature toxic BCW protein having the corresponding amino acid sequences shown as residues 28-618 of SEQ ID NO: 2 or residues 29-619 SEQ ID NO: 4 or 6.

[0073] An embodiment of the invention includes recombinant polynucleotide compositions that encode toxic BCW proteins. For example, toxic BCW proteins can be expressed using recombinant DNA constructs in which an isolated polynucleotide molecule with an open reading frame encoding the protein is operably linked to elements such as a promoter and any other regulatory element that functions to be expressed in the system for which the construct is intended. . Reference in this application to "an isolated DNA molecule", or an equivalent term or phrase, is intended to mean that a DNA molecule is a molecule that is present alone or in combination with other compounds, but not in its natural environment. For example, nucleic acid elements, such as a coding sequence, an intron sequence, an untranslated leader sequence, a promoter sequence, a transcription termination sequence, and the like, that are naturally found in the DNA of an organism's genome are not considered "isolated" as long as the element is found in the organism's genome and at the location in the genome where it is naturally found. However, each of these elements, and portions of such elements, will be "isolated" within the scope of this disclosure if the element is not in the organism's genome, nor the location in the genome where it is naturally found. Similarly, a nucleotide sequence encoding an insecticidal protein, or any naturally occurring insecticidal variant of such a protein, will be an isolated nucleotide sequence, as long as the nucleotide sequence is not in the DNA of a bacterium in which the sequence encoding the protein is naturally found. A synthetic nucleotide sequence encoding the amino acid sequence of a naturally occurring insecticidal protein will be considered isolated for the purposes of this disclosure. For the purposes of this disclosure, any transgenic nucleotide sequence, i.e. a DNA nucleotide sequence inserted into the genome of plant or bacterial cells, or present in an extrachromosomal vector, will be considered an isolated nucleotide sequence, whether it is present in a plasmid or similar structure, used for cell transformation, in the genome of a plant or bacterium, or present in detectable amounts in tissues, progeny, biological samples or commercial products derived from a plant or bacterium. Non-limiting examples include plant-functional promoters operably linked to toxic BCW protein coding sequences for protein expression in plants, or Bt-functional promoters operably linked to BCW toxic protein coding sequences for protein expression in Bt. Other elements that may be operably linked to sequences encoding the toxic BCW protein include, but are not limited to: enhancers, introns, leader sequences, encoded protein immobilization tags (HIS tags), encoded subcellular translocation peptides (e.g., plastid transit peptides, signal peptides), encoded polypeptide sites of post-translational modification enzymes, ribosomal binding sites and RNAi target sites.

[0074] As used herein, the term "recombinant DNA molecule" is a DNA molecule containing a combination of DNA molecules that would not naturally occur together without human intervention. For example, a recombinant DNA molecule may be a DNA molecule that consists of at least two DNA molecules that are heterologous to each other, a DNA molecule that contains a DNA sequence that differs from naturally occurring DNA sequences, or a DNA molecule that that has been incorporated into the DNA of the host cell by genetic transformation or gene editing. Similarly, a "recombinant protein molecule" is a protein molecule containing a combination of amino acids that would not naturally occur together without human intervention. For example, a recombinant protein molecule may be a protein molecule that consists of at least two amino acid molecules that are heterologous to each other, a protein molecule that contains an amino acid sequence that differs from naturally occurring amino acid sequences, or a protein molecule that expressed in the host cell as a result of genetic transformation of the host cell or due to gene editing of the host cell's genome.

[0075] Illustrative recombinant polynucleotide molecules provided herein include, but are not limited to: SEQ ID NO: 7, SEQ ID NO: 9, and SEQ ID NO: 11, as well as each of the nucleotide segments listed in SEQ ID NO: 3 and SEQ ID NO: 5 and which encode the corresponding polypeptides or proteins having the amino acid sequence as indicated in SEQ ID NO: 2 (BCW oo1), SEQ ID NO: 4 (BCW 002), SEQ ID NO: 6 (BCW 003) and SEQ ID NO: 8, SEQ ID NO: 10, and SEQ ID NO: 12. The codons of a recombinant polynucleotide molecule encoding proteins disclosed herein may be replaced with synonymous codons (also referred to as silent substitution). Also provided are recombinant polynucleotides encoding any of the BCW toxin variant proteins described herein.

[0076] A recombinant DNA construct containing sequences encoding a toxic BCW protein may also further comprise a DNA region that encodes one or more insect inhibitory agents that can be configured to co-express or co-express with a DNA sequence encoding a toxic a BCW protein, a protein other than a toxic BCW protein, an insect-inhibiting dsRNA molecule, or an insecticidal chemical. Non-limiting examples of insecticidal chemicals are organochlorides, organophosphates and carbamates, pyrethroids, neonicotinoids and ryanoids.

[0077] The recombinant DNA construct can be assembled such that all proteins or dsRNA molecules are expressed by a single promoter, or each protein or dsRNA molecule is under the control of a single promoter or some combination thereof. The proteins disclosed herein may be expressed from a multigene expression system in which one or more of the proteins described herein are expressed from a common nucleotide segment that also contains other open reading frames and/or promoters depending on the type of expression system chosen. . For example, a bacterial multigene expression system may use a single promoter to drive the expression of multiple/tandem open reading frames from a single operon. In another example, multi-linked expression cassettes can be used in a plant multigene expression system, each expressing a different protein or different agent, such as one or more dsRNA molecules. In yet another example, a plant multigene expression system may use multi-unrelated expression cassettes, each expressing a different protein or different agent, such as one or more dsRNA molecules. A promoter for use in the recombinant nucleic acid described herein may comprise the entire sequence of the promoter, or any variant or fragment thereof having promoter activity or gene regulation activity.

[0078] A recombinant polynucleotide or recombinant DNA construct containing a sequence encoding a toxic BCW protein can be delivered to host cells by vectors, e.g., plasmid, baculovirus, artificial chromosome, virion, cosmid, phagemid, phage, or viral vector. Such vectors can be used to achieve stable or transient expression of a sequence encoding a toxic BCW protein in a host cell, or subsequent expression into a polypeptide. An exogenous construct of a recombinant polynucleotide or recombinant DNA that contains a sequence encoding a toxic BCW protein and that is introduced into a host cell is referred to herein as a "transgene".

[0079] Also provided herein are transgenic bacteria, transgenic plant cells, transgenic plants, fungi and yeasts, and transgenic plant parts that contain any recombinant polynucleotide that expresses any one or more sequences encoding the toxic BCW protein provided herein. . The term "bacterial cell" or "bacteria" may include, but is not limited to, an Agrobacterium, Bacillus, Escherichia, Salmonella, Pseudomonas, or Rhizobium cell. The term "plant cell" or "plant" may include, but is not limited to, a plant cell or plant: alfalfa, banana, barley, beans, broccoli, cabbage, ornamental cabbage, carrots, cassava, castor beans, cauliflower, celery, chickpeas , Chinese cabbage, citrus, coconut, coffee, corn, clover, cotton, cucurbits, cucumber, Menzies pseudo-hemzies, eggplant, eucalyptus, flax, garlic, grapes, hops, leek, lettuce, frankincense pine, millet, melon , walnut, oats, olive tree, onion, ornamental plants, palm, pasture grasses, peas, peanuts, peppers, pigeon peas, pine, potatoes, poplars, pumpkins, radish pine, radish, rapeseed, rice, rhizomes, rye, wild saffron, shrubby, sorghum, southern pine, soybeans, spinach, cucurbits, strawberries, sugar beet, sugar cane, sunflower, sweet corn, amber, sweet potato, switchgrass, tea, tobacco, tomato, triticale, turfgrass, watermelon and wheat . In some embodiments, transgenic plants and transgenic plant parts regenerated from cells of the transgenic plant are provided. In some embodiments, transgenic plants can be obtained from transgenic seed by propagating, cutting, breaking off, chopping, or otherwise detaching a portion from the plant. In some embodiments, the plant part may be a seed, pod, leaf, flower, stem, root, or any part thereof, or a non-regenerated fragment of a part of a transgenic plant. As used herein, a "non-regenerating" portion of a transgenic plant part is a portion that cannot be stimulated to form a whole plant, or that cannot be stimulated to form a whole plant capable of sexual and/or asexual reproduction. In some embodiments, the non-regenerated portion of the plant part is a portion of a transgenic seed, pod, leaf, flower, stem, or root.

[0080] This document also provides methods for producing transgenic plants that contain an insect or lepidoptera inhibitory amount of a toxic BCW protein. Such plants can be obtained by introducing a recombinant polynucleotide that encodes any of the toxic BCW proteins provided herein into a plant cell and selecting a plant derived from said plant cell that expresses an insect or lepidoptera inhibitory amount of the toxic BCW protein. Plants can be obtained from plant cells by regeneration methods, seed, pollen or meristem transformation methods.

[0081] Methods for transforming y plants are known in the art. For example, Agrobacterium-mediated transformation is described in US Patent Application Publication No. 2009/0138985A1 (soybean), No. 2008/0280361A1 (soybean), No. 2009/0142837A1 (corn), No. 2008/0282432 (cotton), No. 2008/0256667 ( cotton), US Pat. No. 2003/0110531 (wheat), US Pat. No. 2001/0042257 A1 (sugar beet), US Pat. (1998) Transgenic Res. 7:213-222 (sugar cane).

[0082] This document also provides the use of a transgenic plant that expresses an insect or lepidopteran inhibitory amount of a toxic BCW protein to control insect or lepidoptera infestation. Any of the aforementioned transgenic plants can be used in the methods for protecting a plant from insect or lepidopteran infestation provided herein. Methods for obtaining transgenic plants that express proteins active against Lepidoptera, such as Cry1A (US patent No. 5880275), Cry1B (US patent application No. 10/525318), Cry1C (US patent No. 6033874), Cry1F proteins, Cry1A/F chimeras ( US Pat.

[0083] This document also proposes the use of any of the above transgenic host cells for the production of toxic BCW protein.

[0084] Additional aspects of the invention include antibodies and methods for detecting polynucleotides that encode for toxic BCW proteins, or methods for distinguishing fragments and segments thereof, methods for identifying additional members of the insect inhibitory protein family, formulations, and methods for controlling insect growth and/or infestation. , and also methods for providing such control to plants and other recipient hosts.

[0085] In some embodiments, the plant product may include commercial or other commercial products derived from the transgenic plant or part of the transgenic plant, wherein the commercial or other products can be tracked commercially by detecting nucleotide segments, or expressed RNA or proteins, that encode or contain distinctive portions of the toxic BCW protein. Such commercial or other commercial products include, but are not limited to: plant parts, biomass, oil, meal, sugar, animal feed, flour, flakes, bran, fluff, processed seeds and seeds.

[0086] Also provided are processed plant products, said processed product containing a detectable amount of a recombinant polynucleotide encoding a toxic BCW protein, a segment thereof, an insect-inhibiting fragment thereof, or any distinctive portion thereof. In some embodiments, the processed product is selected from the group consisting of: vegetable biomass, oil, meal, animal feed, flour, flakes, bran, fluff, husks, and processed seeds. In some embodiments, the processed product is non-recyclable.

[0087] Also provided herein are methods for controlling insects. In some embodiments, Lepidoptera infestation of crop plants is controlled. Such methods may include growing a plant containing an insect or lepidoptera inhibitory amount of the toxic BCW protein. In certain embodiments, such methods may further include any one or more of: (i) applying any composition containing or encoding a toxic BCW protein to a plant or seed that produces a plant; and/or (ii) transforming a plant or plant cell that produces a plant with a polynucleotide encoding a toxic BCW protein. In some embodiments, the plant is a transiently or stably transformed transgenic plant containing a transgene that expresses an insect or lepidoptera inhibitory amount of the toxic BCW protein. In some embodiments, the plant is a non-transgenic plant that has been coated with a composition containing the toxic BCW protein. In some embodiments of such methods, the plant is a corn or sugarcane plant. In some embodiments, the Lepidoptera species is Agrotis ipsilon. In some embodiments, the Lepidoptera species is Diatraea saccharalis. In some embodiments, the Lepidoptera species is in a crop field.

[0088] Increasing the amount of proteins disclosed herein, either in plants or by a process, may include culturing recombinant Bt cells under conditions to express/produce the recombinant polypeptide/proteins. Such a process may include obtaining by drying, lyophilization, homogenization, extraction, filtration, centrifugation, sedimentation or concentration of a culture of recombinant Bt cells expressing/producing the specified recombinant polypeptide. Such a process may result in a Bt cell extract, cell suspension, cell homogenate, cell lysate, cell supernatant, cell filtrate, or cell pellet. By obtaining recombinant polypeptides/proteins thus produced, a composition that contains recombinant polypeptides/proteins may contain bacterial cells, bacterial spores and parasporal intracellular bodies, and can be prepared for various applications, including insect-inhibiting sprayable agricultural products, or in the form formulations that suppress insects in food bioassays.

[0089] In one embodiment, the insect-inhibiting composition/formulation comprising the disclosed recombinant polypeptide/protein may further comprise at least one additional polypeptide that exhibits inhibitory activity against the same Lepidoptera insect species, but differs from a recombinant polypeptide to reduce the occurrence of resistance in Lepidoptera insects to the toxic BCW protein or other Lepidoptera-inhibiting composition. Such a polypeptide is selected from the group consisting of an insect-inhibiting protein, an insect-inhibiting dsRNA molecule, and a chemical compound. One example of the use of such ribonucleotide sequences for insect pest control is described in US Patent Application Publication No. 2006/0021087. Examples of other such compositions include, but are not limited to: Cry1A (US patent No. 5880275), Cry1Ab, Cry1Ac, Cry1Ae, Cry1B (US patent application No. 10/525318), Cry1C (US patent No. 6033874), Cry1E, Cry1F and Cry1A/F chimeras (US Pat. No. 7070982; No. 6962705 and No. 6713063), Cry1G, Cry1H, Cry1I, Cry1J, Cry1K, Cry1L, Cry2A, Cry2Ab (US Pat. No. 7064249), Cry2Ae, Cry4B, Cry6, Cry7, Cry8 , Cry9, Cry15, Cry43A, Cry43B, ET35, ET66, TIC400, TIC800, TIC807, TIC834, TIC853 and TIC1415. Other non-limiting examples are VIP, Axmi and DIG active against Lepidoptera, such as but not limited to Vip3A, VIP3Ab, AXMI-184, AXMI-196, DIG-3, DIG-4, DIG-5 and DIG-11, which can be combined with the proteins described in this document.

[0090] In other embodiments, such a composition may further comprise at least one additional polypeptide that exhibits insect inhibitory activity that is not inhibited by another toxic insect inhibitory BCW protein to broaden the insect inhibition spectrum obtained. For example, combinations of insect-inhibiting toxic BCW proteins with beetle-active proteins such as, but not limited to, Cry1C variants, Cry3A variants, Cry3Bb (US Pat. No. 6,501,009), Cry34 can be used to control beetle insect pests. /35, 5307, Axmi184, Axmi205, AxmiR1, TIC407, TIC417, TIC431, TIC901, TIC1201, TIC3131, DIG-10, and eHIP (U.S. Patent Application Publication No. 2010/0017914).

[0091] The ability of insects to develop resistance to certain insecticides has been documented in the art. One strategy for controlling insect resistance is to use transgenic crops that express two different insect inhibitory agents that have different modes of action. Therefore, any insect that is resistant to one of the insect inhibitory agents can be controlled by the other insect inhibitory agent. Another insect resistance control strategy involves using plants that are not immune to Lepidoptera pest species to create an ecological refuge for them. One specific example is described in US Pat. No. 6,551,962.

[0092] Other embodiments disclosed herein include topical pesticidal chemicals that are designed to control pests that are also controlled by the proteins disclosed herein, for use with proteins in seed treatment, spraying, irrigated, or irrigated with formulations that can be applied directly to the soil (soil irrigation) applied to cultivated plants expressing the proteins disclosed herein, or prepared for application to seeds containing one or more transgenes encoding one or more open proteins. Such formulations for use in seed treatment can be applied with various adhesives and tackifiers known in the art. Such formulations may contain pesticides that are synergistic in RD with the disclosed proteins, such that the pesticide formulation has a different RD for controlling the same or similar pests that may be controlled by the disclosed proteins, or such that the effects of such pesticides control a wider range of pest hosts such as Lepidoptera or Hemiptera species (Hemiptera) or other plant pest species such as Coleoptera species (Coleoptera) that are not effectively controlled.

[0093] The above composition/formulation may further comprise an agriculturally acceptable carrier such as bait, powder, dust, pellets, granules, spray, emulsion, colloidal suspension, aqueous solution, Bacillus spore/crystal preparation, seed fertilizer, a recombinant plant cell/plant tissue/seed/plant transformed to express one or more proteins, or a bacterium transformed to express one or more proteins. Depending on the level of insect inhibition or insecticidal inhibition inherent in the recombinant polypeptide and the activity of the formulation to be used in plant or food bioassays, the composition/formulation may contain varying weight amounts of the recombinant polypeptide, for example, from 0.0001% to 0.001% , up to 0.01%, up to 1%, up to 99% by weight of the recombinant polypeptide.

[0094] The proteins disclosed herein can be combined in formulations for topical application to plant canopy, in soil, in formulations for seed treatment, and in formulations with other agents toxic to the target lepidoptera pest species. Such agents include, but are not limited to, Cry1A proteins, Cry1B, Cry1C, Cry1F, Cry1A/F chimeras, and Cry2Ab protein.

EXAMPLES

[0095] The following disclosed embodiments simply represent the invention, which may be embodied in various forms. Thus, the specific structural and functional details disclosed herein should not be interpreted as limiting. It is to be understood that the full disclosure of each source cited herein is incorporated herein by reference.

Example 1

[0096] This example describes the discovery and analysis of the toxic protein BCW 001 and the construction of chimeric toxins BCW 002 and BCW 003.

[0097] Bt strain EG4384 was found to have activity against Lepidoptera in food bioassays using crystal spore preparations. The genome sequence of this strain was obtained, raw read sequences were processed, contigs were assembled from the processed read sequences, open reading frames were identified that showed homology with Cry1 proteins, and the resulting amino acid sequences were analyzed. A particular open reading frame has been identified as set forth in SEQ ID NO: 1 that encodes a derived protein amino acid sequence (BCW 001, SEQ ID NO: 2) that exhibits a novel amino acid sequence compared to most of the Cry1 proteins known in the art. . The resulting open reading frame protein has all the characteristics of a novel Cry1 type protein as it is 1180 amino acids long, and the alignment with known Cry1 proteins indicates that the protein has a characteristic three-domain structure in the range of approximately 600-630 amino-terminal amino acids and a characteristic amino acid structure. Cry1A type protoxin sequences. The polynucleotide sequence encoding this predicted amino acid sequence contains an open reading frame that is also characteristic of Cry1, namely, an NheI restriction site within the DNA segment encoding the C-terminal region of the predicted toxin domain I, and a KpnI restriction site within the DNA segment, encoding the N-terminal part of the predicted prototoxin domain.

[0098] Comparison of the amino acid sequence of BCW 001 toxin with Cry1Ac shows that the amino acid segment corresponding to domain I (amino acids from about position 1 to about position 258) shows only about 67% identity to the same segment in Cry1Ac, the amino acid segment corresponding to domain II (amino acids from about position 58 to about position 460) shows a very low percentage identity with the same segment in Cry1Ac, and the amino acid segment corresponding to domain III (amino acids from about position 460 to about position 607) shows about 63% identity with the domain segment III from Cry1Ah2.

[0099] The DNA segment encoding substantially the predicted domains II and III was excised at the restriction sites NheI to KpnI, and replaced with the corresponding segment of the Cry1Ac coding sequence in an expression vector containing the DNA segment encoding Cry1Ac, resulting in an open reading frame of from, and in-frame connection in sequential order from 5' end to 3' end, the first segment encoding domain I of Cry1Ac, the second segment encoding domain II and III of BCW 001, and the third segment encoding the prototoxin domain of the toxic Cry1Ac protein. This chimeric construct (SEQ ID NO: 3) encodes a chimeric toxic protein referred to herein as BCW 002 (SEQ ID NO: 4). A slight shift in the breakpoint between domain I and domain II results in an open reading frame (SEQ ID NO: 5) encoding a chimeric toxic protein, referred to herein as BCW 003 (SEQ ID NO: 6), having an amino acid sequence that differs from BCW002 only in amino acid position 259. BCW 003, like BCW 001, contains threonine (T) at position 259, and BCW 002 contains isoleucine (I) at this position. BCW 001 differs from BCW 002 and BCW 003 mainly in domain I of the toxin, i.e. amino acids 1-202, while BCW 002 and BCW 002 are almost identical except for the I/T difference at position 259.

Example 2

[00100] This example demonstrates the effectiveness of the biological activity of the BCW 001, 002 and 003 proteins to control lepidoptera pests.

[00101] Transformation of constructs expressing the toxic BCW 001, 002 and 003 proteins into E. coli or into applicable Bacillus thuringiensis or other Bacilli allowed the expressed proteins to be tested in a bioassay and compared with proteins known in the art to be toxic to upsilon scoops such as Cry1Fa and Cry1Ac. The resulting recombinant strains were observed to express a recombinant protein with activity against Lepidoptera pests. The activity in the bioassay was particularly strong when tested against the larvae of the cutworm (Agrotis ipsilon) and Helicoverpa zea. As noted above in the DETAILED DESCRIPTION, BACKGROUND OF THE INVENTION and SUMMARY, very few toxic proteins have been found that exhibit any appreciable level of biological activity against upsilon cutworms, and therefore there is a need in the art to identify such proteins. for use in plants, to protect such plants from bollworm infestation, and to ensure that there is a sufficient supply of additional anti-both bollworm active proteins available to overcome any development of resistance to any such bollworm active proteins, currently in use, such as the toxic Cry1Fa proteins.

Example 3

[00102] This example demonstrates that BCW 001 domains II and III are sufficient to confer biological activity against upsilon cutworms to other toxic Cry1 proteins when such domains replace the corresponding domains in such other Cry1 toxins.

[00103] Many chimeric BCW toxins have been identified with activity against Lepidoptera, in particular two chimeras showed strong activity against BCW, WBC and SCB.

[00104] Constructs with nucleotide sequences encoding Cry1Ab, Cry1Ac, and Cry1Ca were used to construct chimeras containing BCW 001 domain II and domain III segments replacing the corresponding Cry1Ab, Cry1Ac, and Cry1Ca domains, and the resulting native and chimeric proteins were tested in crystal bioassays dispute. Activities against BCW, FAW and CEW in these food bioassays were tabulated. Under experimental testing conditions, Cry1Ac was active against FAW, BCW and CEW, Cry1Ab was active against FAW and CEW but not against BCW, and Cry1Ca was not active against FAW, BCW and also CEW. BCW 001 was active against BCW and CEW, but not FAW. Compared to BCW 003, the activity of Cry1Ac against BCW was approximately ten times less for Cry1Ac. Cry1Ab and Cry1Ac, chimeras containing BCW001 domains II and III, exhibited toxic properties when tested in bioassays against FAW, BCW and CEW. Cry1Ac was not toxic against CEW and Cry1Ab was not toxic against BCW. Cry1Ca/BCW 001 chimeras were constructed in which the Cry1Ca domain III was replaced by the corresponding BCW 001 domain and the resulting chimeric toxin was toxic against FAW, BCW and CEW, while the Cry1Ca toxin was ineffective when tested against any of these pests.

Example 4

[00105] This example illustrates the toxic properties of BCW 001, 002 and 003 when tested in a bioassay against a variety of Lepidoptera pests.

[00106] Protocols for bioanalysis and scoring of insect mortality and stunting are known in the art, examples of which are described in PCT Patent Application Publication No. WO 2012/139004 and US Pat. No. 7,927,598.

[00107] In FIG. 1 correlates various toxic proteins BCW 001, 002 and 003 for pesticidal activity to insect species in food bioassays. Each of these toxic proteins has shown activity against Lepidoptera.

Example 5

[00108] This example describes the construction of artificial sequences encoding the proteins of this invention for use in plants, the preparation of plant vectors and constructs for use in plants, and the production of plants expressing the proteins of this invention.

[00109] Nucleotide sequences encoding BCW 001 protein (SEQ ID NO: 1), BCW 002 protein (SEQ ID NO: 3), and BCW 003 protein (SEQ ID NO: 5) were designed and synthesized according to the methods described in US Pat. No. 5,500,365. These engineered coding regions for expression in plants are shown herein as SEQ ID NO: 7 encoding BCW 001, SEQ ID NO: 9 encoding BCW 002, and SEQ ID NO: 11 encoding BCW 003.

[00110] A variety of plant expression cassettes were constructed with the sequences set forth in SEQ ID NOs: 7, 9, and 11. Such expression cassettes are useful for transient expression in plant protoplasts or for plant cell transformation. Exemplary expression cassettes have been designed to accommodate the possible placement of the protein in the cell. One set of expression cassettes was designed to allow the protein to be translated and remain in the cytosol. Another set of expression cassettes was designed to have a transit peptide adjacent to a toxic protein to allow targeting of a cell organelle such as a chloroplast or plastid. All expression cassettes were designed to start at the 5' end with a promoter, which may consist of several promoter elements and enhancer elements contiguously linked to enhance expression of the transgene. The promoter sequence was typically followed contiguous by one or more leader sequences 3' to the promoter. The intron sequence was placed 3' to the leader sequence to improve transgene expression. The coding sequence for the toxin or transit peptide and the coding sequence for the toxin were located 3' to the promoter, leader and intron. A specific sequence has been placed 3' to the coding sequence to facilitate termination of transcription and provides sequences important for polyadenylation of the resulting transcript. All of the elements described above were contiguous, often with additional sequence provided to construct the expression cassette, such as endonuclease restriction sites or non-ligase cloning sites.

[00111] For maize plants, a set of expression cassettes for cytosolic expression BCW 001 containing the Mexicana ubiquitin 1 promoter, BCW 002 containing the Orysza sativa actin 15 promoter or 35S promoter, and BCW 003 containing the 35S promoter was developed.

[00112] Another set of expression cassettes was designed to target the expression in maize plants of the insect-toxic proteins BCW 002 and BCW 003, in which the sequence encoding a chloroplast peptide (e.g., CTP2) was fused in-frame at the 5' end of the DNA segment encoding toxic BCW proteins containing the Orysza sativa actin 15 promoter or the 35S promoter and a sequence containing the 35S promoter.

[00113] Sugar cane expression cassettes containing the CaMV 35S promoter or the PClSV.FLt promoter operably linked to the 35S promoter were constructed as plant transformation vectors. In some cases, a chloroplast-targeted Cry2Ab expression cassette was included.

[00114] Plants expressing the proteins of this invention were tested against third molt BCW, WBC, CEW, SWC and SCB larvae. The cytosol expression cassette for BCW 001, and the cytosol- and plastid-targeted expression cassette for BCW 002 and BCW 003 were cloned and used to generate transgenic corn plants expressing these proteins. Transformed cells were induced to form plants by methods known in the art. Bioassay using plant leaf discs was performed in a manner similar to that described in US Pat. No. 8,344,207. Leaf damage rate (LDR) was assigned a score based on the percentage of leaf disc eaten by the insect, on a scale from 0 (0% eaten) to 11 (more than fifty%). The degree estimation step is increased by 5%. An isogenic maize line was used to obtain tissue as a negative control and the results were evaluated. Both expression in target plastids and cytosolic expression of insect-toxic proteins BCW 002 and BCW 003 reduced nutritional damage compared to non-transformed controls. The results of leaf disc analyzes for these insects are consistent with the bioassay data from the examples above. One construct containing the BCW 003 cassette for cytosolic expression produced 34 transformants, 25 of which showed complete control of BCW larvae. Maize plants expressing BCW 001 and BCW 003 in the cytosol were also tested for CEW, SWC and FAW. Plants expressing BCW 003 showed 100% control of CEW and SWC and LDR values ranging from 1 to 2. The three transformants expressing BCW 001 gave rise to plants showing efficacy against CEW and SWC and LDR values ranging from 1 to 2. 3. This is consistent with the food bioassay data presented in the previous examples.

[00115] Transgenic sugar cane plants expressing BCW 003 were generated and tested against SCB in bioassays. Each bioassay included leaf discs from wild-type sugarcane as a negative control, and from a positive control expressing high levels of Cry2Ab. Insect mortality and leaf damage were measured four (4) days after infection. Leaf discs of several sugar cane transformants expressing BCW 003 were found to show control of Diatraea saccharalis (sugarcane borer) in planta and have a damage score below 2, similar to the positive control, and an average insect mortality of 90-100%.

[00116] Transformants expressing BCW 003 and Cry2Ab showed better control of SCB compared to transformants that expressed only BCW 003.

--->

SEQUENCE LIST

<110> Monsanto Technology LLC

<120> PESTICIDAL PROTEIN TOXINS ACTIVE AGAINST

Lepidoptera

<130> MONS:434WO

<150> US 62/445,313

<151> 2017-01-12

<160> 12

<170> PatentIn version 3.5

<210> 1

<211> 3543

<212> DNA

<213> Bacillus thuringiensis

<220>

<221> CDS

<222> (1)..(3540)

<400> 1

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Met Glu Glu Asn Asn Gln Asn Gln Cys Val Pro Tyr Asn Cys Leu Asn

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aat cct gca atc gaa ata tta gaa gga gac aga ata tca gtt ggt aac 96

Asn Pro Ala Ile Glu Ile Leu Glu Gly Asp Arg Ile Ser Val Gly Asn

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act cca atc gat att tct cta tca ctt gtg gaa ctt ctt att agt gaa 144

Thr Pro Ile Asp Ile Ser Leu Ser Leu Val Glu Leu Leu Ile Ser Glu

35 40 45

ttt gtc cca ggc ggt gga ata ata aca gga ttg ttg aac ata gta tgg 192

Phe Val Pro Gly Gly Gly Ile Ile Thr Gly Leu Leu Asn Ile Val Trp

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gga ttt gta ggg cct tcc caa tgg gac gca ttt ctt gct caa gtg gaa 240

Gly Phe Val Gly Pro Ser Gln Trp Asp Ala Phe Leu Ala Gln Val Glu

65 70 75 80

cag tta att aac caa agg ata tca gaa gct gta aga aat aca gca att 288

Gln Leu Ile Asn Gln Arg Ile Ser Glu Ala Val Arg Asn Thr Ala Ile

85 90 95

cag gaa tta gag gga atg gcg cgg gtt tat aga acc tat gct act gct 336

Gln Glu Leu Glu Gly Met Ala Arg Val Tyr Arg Thr Tyr Ala Thr Ala

100 105 110

ttt gct gag tgg gaa aga gat cct aat aac aca gat cta aga gaa gca 384

Phe Ala Glu Trp Glu Arg Asp Pro Asn Asn Thr Asp Leu Arg Glu Ala

115 120 125

gta cgg aca cag ttt aca gca act gag act tat atc agt gga aga ata 432

Val Arg Thr Gln Phe Thr Ala Thr Glu Thr Tyr Ile Ser Gly Arg Ile

130 135 140

tct gtt tta aaa att caa aat ttt gaa gtg cag ctg tta tcg gtg ttt 480

Ser Val Leu Lys Ile Gln Asn Phe Glu Val Gln Leu Leu Ser Val Phe

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Ala Gln Ala Ala Asn Leu His Leu Ser Leu Leu Arg Asp Val Val Phe

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Phe Gly Gln Arg Trp Gly Phe Ser Thr Thr Thr Val Asn Asn Tyr Tyr

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aat gat tta aca gaa gag att agt acc tat aca gat tat gca gta cgc 624

Asn Asp Leu Thr Glu Glu Ile Ser Thr Tyr Thr Asp Tyr Ala Val Arg

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tgg tac aat acg gga tta gag cgt gta tgg gga ccg gat tct aga gat 672

Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg Asp

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tgg gta agg tat aat caa ttt aga aga gag cta aca ctt act gta tta 720

Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val Leu

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gat atc gtt gct cta ttc cca aat tat gat agt cga agg tat cca att 768

Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro Ile

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cga aca gtt tcc caa tta aca aga gaa att tat acg aac cca gta tta 816

Arg Thr Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val Leu

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gaa aat ttt gat ggt agt ttt cgt gga atg gct cag aga ata gaa cag 864

Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu Gln

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aat att agg caa cca cat ctt atg gat atc ctt aat agt ata acc att 912

Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr Ile

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tat act gat gtg cat aga ggc ttt aat tat tgg tca ggg cat caa ata 960

Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln Ile

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aca gct tct cct gta ggg ttt tca gga cca gaa ttc gca ttc cct tta 1008

Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro Leu

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ttt ggg aat gcg gga aat gca gct cca ccc gta ctt gtc tca tta act 1056

Phe Gly Asn Ala Gly Asn Ala Ala Pro Pro Val Leu Val Ser Leu Thr

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ggt ttg ggg att ttt aga aca tta tct tca cct tta tat aga aga att 1104

Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg Ile

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Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp Gly

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acg gag ttt tct ttt gcc tcc cta acg acc aac ttg cct tcc act ata 1200

Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr Ile

385 390 395 400

tat aga caa agg ggt aca gtc gat tca cta gat gta ata ccg cca cag 1248

Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro Gln

405 410 415

gat aat agt gta cca cct cgt gcg gga ttt agc cat cga ttg agt cat 1296

Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser His

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gtt aca atg ctg agc caa gca gct gga gca gtt tac acc ttg aga gct 1344

Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg Ala

435 440 445

cca acg ttt tct tgg cag cat cgc agt gct acg aca act aat ata att 1392

Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile Ile

450 455 460

gca gcg gat agt att act caa att cct gct gtt aaa gga cgt tct att 1440

Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser Ile

465 470 475 480

att aat aat ggc acg gta att tca gga cca ggg ttt acc gga ggc gat 1488

Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly Asp

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ttg gtt aga tta tac aat gct gat ttt aat att aat aat aga gca tac 1536

Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala Tyr

500 505 510

ctt gaa gtt ccg ata ttc ttc caa tca ccc tct aca aat tat cgt gtt 1584

Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg Val

515 520 525

cgt gtt cgt tat gct tct aca tct tca ctc cct gta gat gta gtt ttc 1632

Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val Phe

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gga aat att agt cat cct act aca ttc cca gcc act gcc aga tca tta 1680

Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser Leu

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gat aat cta caa tcc aat gat ttt gga tat att gat att gct gga act 1728

Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly Thr

565 570 575

ttc tta cct tca cta ggg cct agt ata ggt atc aga cct atg tta tct 1776

Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu Ser

580 585 590

act att aat ttg ata gta gat aga ttt gaa ttt att cca gta act gca 1824

Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr Ala

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acc ttt gaa gca gaa tcg gat tta gaa aga gca caa aag gcg gtg aat 1872

Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val Asn

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gcg ctg ttt act tct aca aac caa cta ggg ata aaa aca gat gtg acg 1920

Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val Thr

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Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser Asp

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gaa ttt tat ctg gat gaa aag cga gaa ttg tcc gag aaa gtc aaa cat 2016

Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys His

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gcg aag cga ctc agt gat gag cga aat tta ctt caa gat cca aac ttc 2064

Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn Phe

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agg ggc atc aat aga caa cca gat cgt ggc tgg aga gga agt acg gat 2112

Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr Asp

690 695 700

att acc atc caa gga gga gat gac gta ttc aaa gag aat tac gtc aca 2160

Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val Thr

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cta cca ggt acc ttt gat gag tgc tat cca acg tat tta tat caa aaa 2208

Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln Lys

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ata gat gag tcg aaa tta aaa gcc tat acc cgt tat caa tta aga ggg 2256

Ile Asp Glu Ser Lys Leu Lys Ala Tyr Thr Arg Tyr Gln Leu Arg Gly

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Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr Asn

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Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp Pro

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Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg Cys

785 790 795 800

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Ala Pro His Leu Glu Trp Asn Pro Asp Leu His Cys Ser Cys Arg Asp

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ggg gaa aaa tgt gct cat cat tct cat cat ttc tcc ttg gac att gat 2496

Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile Asp

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gtt gga tgt aca gac tta aat gag gat tta ggt gta tgg gtg ata ttc 2544

Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile Phe

835 840 845

aag att aag acg caa gat ggc cat gca aga cta gga aat cta gag ttt 2592

Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu Phe

850 855 860

ctc gaa gag aaa cca tta gta ggg gaa gca cta gct cgt gtg aaa aga 2640

Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys Arg

865 870 875 880

gcg gag aaa aaa tgg aga gac aaa cgc gaa aaa tta caa ttg gaa aca 2688

Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Gln Leu Glu Thr

885 890 895

aat atc gtt tat aaa gag gca aaa gaa tct gta gat gct tta ttt gta 2736

Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe Val

900 905 910

aac tct caa tat gat caa tta caa gcg gat acg aac atc gcg atg att 2784

Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met Ile

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His Ala Ala Asp Lys Arg Val His Arg Ile Arg Glu Ala Tyr Leu Pro

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gag tta tct gtg att ccg ggt gta aat gca gac att tcc gaa gaa tta 2880

Glu Leu Ser Val Ile Pro Gly Val Asn Ala Asp Ile Ser Glu Glu Leu

945 950 955 960

gaa ggg cgt att ttc act gca ttc tct cta tat gat gcg aga aat gtc 2928

Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn Val

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att aaa aat ggc gat ttc aat aat ggc tta tta tgc tgg aac gtg aaa 2976

Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Leu Cys Trp Asn Val Lys

980 985 990

ggg cat gta gat gta gaa gaa caa aat aac cac cgt tcg gtc ctt gtt 3024

Gly His Val Asp Val Glu Glu Gln Asn Asn His Arg Ser Val Leu Val

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Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val Cys

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ccg ggg cgt ggc tat atc ctt cgt gtc aca gcg tac aag gag gga 3114

Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu Gly

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tat gga gaa ggt tgc gta acc att cat gag atc gag aac aat aca 3159

Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn Thr

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Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Val Tyr Pro

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Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Ala Asn Gln Glu Glu

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tat gag ggt acg tac act tct cgt aat cga gga tat gac gaa gcc 3294

Tyr Glu Gly Thr Tyr Thr Ser Arg Asn Arg Gly Tyr Asp Glu Ala

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Tyr Glu Ser Asn Ser Ser Val Pro Ala Glu Tyr Ala Ser Val Tyr

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gaa gaa aaa gtg tat aca gat gga cga aga ggg aat cct tgt gaa 3384

Glu Glu Lys Val Tyr Thr Asp Gly Arg Arg Gly Asn Pro Cys Glu

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tct aac aga gga tat ggg gat tac aca cca cta cca gct ggc tat 3429

Ser Asn Arg Gly Tyr Gly Asp Tyr Thr Pro Leu Pro Ala Gly Tyr

1130 1135 1140

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Val Thr Lys Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp

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att gag att gga gaa aca gaa gga aca ttc att gtg gat agt gtg 3519

Ile Glu Ile Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val

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gaa tta ctc ctt atg gag gaa taa 3543

Glu Leu Leu Leu Met Glu Glu

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<210> 2

<211> 1180

<212> PROTEIN

<213> Bacillus thuringiensis

<400> 2

Met Glu Glu Asn Asn Gln Asn Gln Cys Val Pro Tyr Asn Cys Leu Asn

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Asn Pro Ala Ile Glu Ile Leu Glu Gly Asp Arg Ile Ser Val Gly Asn

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Thr Pro Ile Asp Ile Ser Leu Ser Leu Val Glu Leu Leu Ile Ser Glu

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Phe Val Pro Gly Gly Gly Ile Ile Thr Gly Leu Leu Asn Ile Val Trp

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Gly Phe Val Gly Pro Ser Gln Trp Asp Ala Phe Leu Ala Gln Val Glu

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Gln Leu Ile Asn Gln Arg Ile Ser Glu Ala Val Arg Asn Thr Ala Ile

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Gln Glu Leu Glu Gly Met Ala Arg Val Tyr Arg Thr Tyr Ala Thr Ala

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Phe Ala Glu Trp Glu Arg Asp Pro Asn Asn Thr Asp Leu Arg Glu Ala

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Val Arg Thr Gln Phe Thr Ala Thr Glu Thr Tyr Ile Ser Gly Arg Ile

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Ser Val Leu Lys Ile Gln Asn Phe Glu Val Gln Leu Leu Ser Val Phe

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Ala Gln Ala Ala Asn Leu His Leu Ser Leu Leu Arg Asp Val Val Phe

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Phe Gly Gln Arg Trp Gly Phe Ser Thr Thr Thr Val Asn Asn Tyr Tyr

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Asn Asp Leu Thr Glu Glu Ile Ser Thr Tyr Thr Asp Tyr Ala Val Arg

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Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg Asp

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Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val Leu

225 230 235 240

Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro Ile

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Arg Thr Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val Leu

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Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu Gln

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Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr Ile

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Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln Ile

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Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro Leu

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Phe Gly Asn Ala Gly Asn Ala Ala Pro Pro Val Leu Val Ser Leu Thr

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Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg Ile

355 360 365

Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp Gly

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Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr Ile

385 390 395 400

Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro Gln

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Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser His

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Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg Ala

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Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile Ile

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Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser Ile

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Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly Asp

485 490 495

Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala Tyr

500 505 510

Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg Val

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Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val Phe

530 535 540

Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser Leu

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Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly Thr

565 570 575

Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu Ser

580 585 590

Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr Ala

595 600 605

Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val Asn

610 615 620

Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val Thr

625 630 635 640

Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser Asp

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Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys His

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Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn Phe

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Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr Asp

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Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val Thr

705 710 715 720

Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln Lys

725 730 735

Ile Asp Glu Ser Lys Leu Lys Ala Tyr Thr Arg Tyr Gln Leu Arg Gly

740 745 750

Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr Asn

755 760 765

Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp Pro

770 775 780

Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg Cys

785 790 795 800

Ala Pro His Leu Glu Trp Asn Pro Asp Leu His Cys Ser Cys Arg Asp

805 810 815

Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile Asp

820 825 830

Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile Phe

835 840 845

Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu Phe

850 855 860

Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys Arg

865 870 875 880

Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Gln Leu Glu Thr

885 890 895

Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe Val

900 905 910

Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met Ile

915 920 925

His Ala Ala Asp Lys Arg Val His Arg Ile Arg Glu Ala Tyr Leu Pro

930 935 940

Glu Leu Ser Val Ile Pro Gly Val Asn Ala Asp Ile Ser Glu Glu Leu

945 950 955 960

Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn Val

965 970 975

Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Leu Cys Trp Asn Val Lys

980 985 990

Gly His Val Asp Val Glu Glu Gln Asn Asn His Arg Ser Val Leu Val

995 1000 1005

Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val Cys

1010 1015 1020

Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu Gly

1025 1030 1035

Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn Thr

1040 1045 1050

Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Val Tyr Pro

1055 1060 1065

Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Ala Asn Gln Glu Glu

1070 1075 1080

Tyr Glu Gly Thr Tyr Thr Ser Arg Asn Arg Gly Tyr Asp Glu Ala

1085 1090 1095

Tyr Glu Ser Asn Ser Ser Val Pro Ala Glu Tyr Ala Ser Val Tyr

1100 1105 1110

Glu Glu Lys Val Tyr Thr Asp Gly Arg Arg Gly Asn Pro Cys Glu

1115 1120 1125

Ser Asn Arg Gly Tyr Gly Asp Tyr Thr Pro Leu Pro Ala Gly Tyr

1130 1135 1140

Val Thr Lys Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp

1145 1150 1155

Ile Glu Ile Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val

1160 1165 1170

Glu Leu Leu Leu Met Glu Glu

1175 1180

<210> 3

<211> 3534

<212> DNA

<213> Artificial sequence

<220>

<223> Fully synthetic DNA encoding BCW 002

<220>

<221> CDS

<222> (1)..(3531)

<400> 3

atg gat aac aat ccg aac atc aat gaa tgc att cct tat aat tgt tta 48

Met Asp Asn Asn Pro Asn Ile Asn Glu Cys Ile Pro Tyr Asn Cys Leu

1 5 10 15

agt aac cct gaa gta gaa gta tta ggt gga gaa aga ata gaa act ggt 96

Ser Asn Pro Glu Val Glu Val Leu Gly Gly Glu Arg Ile Glu Thr Gly

20 25 30

tac acc cca atc gat att tcc ttg tcg cta acg caa ttt ctt ttg agt 144

Tyr Thr Pro Ile Asp Ile Ser Leu Ser Leu Thr Gln Phe Leu Leu Ser

35 40 45

gaa ttt gtt ccc ggt gct gga ttt gtg tta gga cta gtt gat ata ata 192

Glu Phe Val Pro Gly Ala Gly Phe Val Leu Gly Leu Val Asp Ile Ile

50 55 60

tgg gga att ttt ggt ccc tct caa tgg gac gca ttt ctt gta caa att 240

Trp Gly Ile Phe Gly Pro Ser Gln Trp Asp Ala Phe Leu Val Gln Ile

65 70 75 80

gaa cag tta att aac caa aga ata gaa gaa ttc gct agg aac caa gcc 288

Glu Gln Leu Ile Asn Gln Arg Ile Glu Glu Phe Ala Arg Asn Gln Ala

85 90 95

att tct aga tta gaa gga cta agc aat ctt tat caa att tac gca gaa 336

Ile Ser Arg Leu Glu Gly Leu Ser Asn Leu Tyr Gln Ile Tyr Ala Glu

100 105 110

tct ttt aga gag tgg gaa gca gat cct act aat cca gca tta aga gaa 384

Ser Phe Arg Glu Trp Glu Ala Asp Pro Thr Asn Pro Ala Leu Arg Glu

115 120 125

gag atg cgt att caa ttc aat gac atg aac agt gcc ctt aca acc gct 432

Glu Met Arg Ile Gln Phe Asn Asp Met Asn Ser Ala Leu Thr Thr Ala

130 135 140

att cct ctt ttg gca gtt caa aat tat caa gtt cct ctt tta tca gta 480

Ile Pro Leu Leu Ala Val Gln Asn Tyr Gln Val Pro Leu Leu Ser Val

145 150 155 160

tat gtt caa gct gca aat tta cat tta tca gtt ttg aga gat gtt tca 528

Tyr Val Gln Ala Ala Asn Leu His Leu Ser Val Leu Arg Asp Val Ser

165 170 175

gtg ttt gga caa agg tgg gga ttt gat gcc gcg act atc aat agt cgt 576

Val Phe Gly Gln Arg Trp Gly Phe Asp Ala Ala Thr Ile Asn Ser Arg

180 185 190

tat aat gat tta act agg ctt att ggc aac tat aca gat tat gct gta 624

Tyr Asn Asp Leu Thr Arg Leu Ile Gly Asn Tyr Thr Asp Tyr Ala Val

195 200 205

cgc tgg tac aat acg gga tta gaa cgt gta tgg gga ccg gat tct aga 672

Arg Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg

210 215 220

gat tgg gta agg tat aat caa ttt aga aga gaa tta aca cta act gta 720

Asp Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val

225 230 235 240

tta gat atc gtt gct ctg ttc ccg aat tat gat agt aga aga tat cca 768

Leu Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro

245 250 255

att cga ata gtt tcc caa tta aca aga gaa att tat acg aac cca gta 816

Ile Arg Ile Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val

260 265 270

tta gaa aat ttt gat ggt agt ttt cgt gga atg gct cag aga ata gaa 864

Leu Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu

275 280 285

cag aat att agg caa cca cat ctt atg gat atc ctt aat agt ata acc 912

Gln Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr

290 295 300

att tat act gat gtg cat aga ggc ttt aat tat tgg tca ggg cat caa 960

Ile Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln

305 310 315 320

ata aca gct tct cct gta ggg ttt tca gga cca gaa ttc gca ttc cct 1008

Ile Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro

325 330 335

tta ttt ggg aat gcg gga aat gca gct cca ccc gta ctt gtc tca tta 1056

Leu Phe Gly Asn Ala Gly Asn Ala Ala Pro Val Leu Val Ser Leu

340 345 350

act ggt ttg ggg att ttt aga aca tta tct tca cct tta tat aga aga 1104

Thr Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg

355 360 365

att ata ctt ggt tca ggc cca aat aat cag gaa ctg ttt gtc ctt gat 1152

Ile Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp

370 375 380

gga acg gag ttt tct ttt gcc tcc cta acg acc aac ttg cct tcc act 1200

Gly Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr

385 390 395 400

ata tat aga caa agg ggt aca gtc gat tca cta gat gta ata ccg cca 1248

Ile Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro

405 410 415

cag gat aat agt gta cca cct cgt gcg gga ttt agc cat cga ttg agt 1296

Gln Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser

420 425 430

cat gtt aca atg ctg agc caa gca gct gga gca gtt tac acc ttg aga 1344

His Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg

435 440 445

gct cca acg ttt tct tgg cat cgc agt gct acg aca act aat ata 1392

Ala Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile

450 455 460

att gca gcg gat agt att act caa att cct gct gtt aaa gga cgt tct 1440

Ile Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser

465 470 475 480

att att aat aat ggc acg gta att tca gga cca ggg ttt acc gga ggc 1488

Ile Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly

485 490 495

gat ttg gtt aga tta tac aat gct gat ttt aat att aat aat aga gca 1536

Asp Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala

500 505 510

tac ctt gaa gtt ccg ata ttc ttc caa tca ccc tct aca aat tat cgt 1584

Tyr Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg

515 520 525

gtt cgt gtt cgt tat gct tct aca tct tca ctc cct gta gat gta gtt 1632

Val Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val

530 535 540

ttc gga aat att agt cat cct act aca ttc cca gcc act gcc aga tca 1680

Phe Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser

545 550 555 560

tta gat aat cta caa tcc aat gat ttt gga tat att gat att gct gga 1728

Leu Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly

565 570 575

act ttc tta cct tca cta ggg cct agt ata ggt atc aga cct atg tta 1776

Thr Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu

580 585 590

tct act att aat ttg ata gta gat aga ttt gaa ttt att cca gta act 1824

Ser Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr

595 600 605

gca acc ttt gaa gca gaa tcg gat tta gaa aga gca caa aag gcg gtg 1872

Ala Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val

610 615 620

aat gcg ctg ttt act tct aca aac caa cta ggg ata aaa aca gat gtg 1920

Asn Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val

625 630 635 640

acg gat tat cat att gat caa gtg tcc aat tta gtg gag tgt tta tcg 1968

Thr Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser

645 650 655

gat gaa ttt tat ctg gat gaa aag cga gaa ttg tcc gag aaa gtc aaa 2016

Asp Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys

660 665 670

cat gcg aag cga ctc agt gat gag cga aat tta ctt caa gat cca aac 2064

His Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn

675 680 685

ttc agg ggc atc aat aga caa cca gat cgt ggc tgg aga gga agt acg 2112

Phe Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr

690 695 700

gat att acc atc caa gga gga gat gac gta ttc aaa gag aat tac gtc 2160

Asp Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val

705 710 715 720

aca cta cca ggt acc ttt gat gag tgc tat cca aca tat ttg tat caa 2208

Thr Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln

725 730 735

aaa atc gat gaa tca aaa tta aaa gcc ttt acc cgt tat caa tta aga 2256

Lys Ile Asp Glu Ser Lys Leu Lys Ala Phe Thr Arg Tyr Gln Leu Arg

740 745 750

ggg tat atc gaa gat agt caa gac tta gaa atc tat tta att cgc tac 2304

Gly Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr

755 760 765

aat gca aaa cat gaa aca gta aat gtg cca ggt acg ggt tcc tta tgg 2352

Asn Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp

770 775 780

ccg ctt tca gcc caa agt cca atc gga aag tgt gga gag ccg aat cga 2400

Pro Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg

785 790 795 800

tgc gcg cca cac ctt gaa tgg aat cct gac tta gat tgt tcg tgt agg 2448

Cys Ala Pro His Leu Glu Trp Asn Pro Asp Leu Asp Cys Ser Cys Arg

805 810 815

gat gga gaa aag tgt gcc cat cat tcg cat cat ttc tcc tta gac att 2496

Asp Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile

820 825 830

gat gta gga tgt aca gac tta aat gag gac cta ggt gta tgg gtg atc 2544

Asp Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile

835 840 845

ttt aag att aag acg caa gat ggg cac gca aga cta ggg aat cta gag 2592

Phe Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu

850 855 860

ttt ctc gaa gag aaa cca tta gta gga gaa gca cta gct cgt gtg aaa 2640

Phe Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys

865 870 875 880

aga gcg gag aaa aaa tgg aga gac aaa cgt gaa aaa ttg gaa tgg gaa 2688

Arg Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Glu Trp Glu

885 890 895

aca aat atc gtt tat aaa gag gca aaa gaa tct gta gat gct tta ttt 2736

Thr Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe

900 905 910

gta aac tct caa tat gat caa tta caa gcg gat acg aat att gcc atg 2784

Val Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met

915 920 925

att cat gcg gca gat aaa cgt gtt cat agc att cga gaa gct tat ctg 2832

Ile His Ala Ala Asp Lys Arg Val His Ser Ile Arg Glu Ala Tyr Leu

930 935 940

cct gag ctg tct gtg att ccg ggt gtc aat gcg gct att ttt gaa gaa 2880

Pro Glu Leu Ser Val Ile Pro Gly Val Asn Ala Ala Ile Phe Glu Glu

945 950 955 960

tta gaa ggg cgt att ttc act gca ttc tcc cta tat gat gcg aga aat 2928

Leu Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn

965 970 975

gtc att aaa aat ggt gat ttt aat aat ggc tta tcc tgc tgg aac gtg 2976

Val Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Ser Cys Trp Asn Val

980 985 990

aaa ggg cat gta gt gta gaa gaa caa aac aac caa cgt tcg gtc ctt 3024

Lys Gly His Val Asp Val Glu Glu Gln Asn Asn Gln Arg Ser Val Leu

995 1000 1005

gtt gtt ccg gaa tgg gaa gca gaa gtg tca caa gaa gtt cgt gtc 3069

Val Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val

1010 1015 1020

tgt ccg ggt cgt ggc tat atc ctt cgt gtc aca gcg tac aag gag 3114

Cys Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu

1025 1030 1035

gga tat gga gaa ggt tgc gta acc att cat gag atc gag aac aat 3159

Gly Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn

1040 1045 1050

aca gac gaa ctg aag ttt agc aac tgc gta gaa gag gaa atc tat 3204

Thr Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Ile Tyr

1055 1060 1065

cca aat aac acg gta acg tgt aat gat tat act gta aat caa gaa 3249

Pro Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Val Asn Gln Glu

1070 1075 1080

gaa tac gga ggt gcg tac act tct cgt aat cga gga tat aac gaa 3294

Glu Tyr Gly Gly Ala Tyr Thr Ser Arg Asn Arg Gly Tyr Asn Glu

1085 1090 1095

gct cct tcc gta cca gct gat tat gcg tca gtc tat gaa gaa aaa 3339

Ala Pro Ser Val Pro Ala Asp Tyr Ala Ser Val Tyr Glu Glu Lys

1100 1105 1110

tcg tat aca gat gga cga aga gag aat cct tgt gaa ttt aac aga 3384

Ser Tyr Thr Asp Gly Arg Arg Glu Asn Pro Cys Glu Phe Asn Arg

1115 1120 1125

ggg tat agg gat tac acg cca cta cca gtt ggt tat gtg aca aaa 3429

Gly Tyr Arg Asp Tyr Thr Pro Leu Pro Val Gly Tyr Val Thr Lys

1130 1135 1140

gaa tta gaa tac ttc cca gaa acc gat aag gta tgg att gag att 3474

Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp Ile Glu Ile

1145 1150 1155

gga gaa acg gaa gga aca ttt atc gtg gac agc gtg gaa tta ctc 3519

Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val Glu Leu Leu

1160 1165 1170

ctt atg gag gaa tag 3534

Leu Met Glu Glu

1175

<210> 4

<211> 1177

<212> PROTEIN

<213> Artificial sequence

<220>

<223> Synthetic construction

<400> 4

Met Asp Asn Asn Pro Asn Ile Asn Glu Cys Ile Pro Tyr Asn Cys Leu

1 5 10 15

Ser Asn Pro Glu Val Glu Val Leu Gly Gly Glu Arg Ile Glu Thr Gly

20 25 30

Tyr Thr Pro Ile Asp Ile Ser Leu Ser Leu Thr Gln Phe Leu Leu Ser

35 40 45

Glu Phe Val Pro Gly Ala Gly Phe Val Leu Gly Leu Val Asp Ile Ile

50 55 60

Trp Gly Ile Phe Gly Pro Ser Gln Trp Asp Ala Phe Leu Val Gln Ile

65 70 75 80

Glu Gln Leu Ile Asn Gln Arg Ile Glu Glu Phe Ala Arg Asn Gln Ala

85 90 95

Ile Ser Arg Leu Glu Gly Leu Ser Asn Leu Tyr Gln Ile Tyr Ala Glu

100 105 110

Ser Phe Arg Glu Trp Glu Ala Asp Pro Thr Asn Pro Ala Leu Arg Glu

115 120 125

Glu Met Arg Ile Gln Phe Asn Asp Met Asn Ser Ala Leu Thr Thr Ala

130 135 140

Ile Pro Leu Leu Ala Val Gln Asn Tyr Gln Val Pro Leu Leu Ser Val

145 150 155 160

Tyr Val Gln Ala Ala Asn Leu His Leu Ser Val Leu Arg Asp Val Ser

165 170 175

Val Phe Gly Gln Arg Trp Gly Phe Asp Ala Ala Thr Ile Asn Ser Arg

180 185 190

Tyr Asn Asp Leu Thr Arg Leu Ile Gly Asn Tyr Thr Asp Tyr Ala Val

195 200 205

Arg Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg

210 215 220

Asp Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val

225 230 235 240

Leu Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro

245 250 255

Ile Arg Ile Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val

260 265 270

Leu Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu

275 280 285

Gln Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr

290 295 300

Ile Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln

305 310 315 320

Ile Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro

325 330 335

Leu Phe Gly Asn Ala Gly Asn Ala Ala Pro Val Leu Val Ser Leu

340 345 350

Thr Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg

355 360 365

Ile Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp

370 375 380

Gly Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr

385 390 395 400

Ile Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro

405 410 415

Gln Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser

420 425 430

His Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg

435 440 445

Ala Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile

450 455 460

Ile Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser

465 470 475 480

Ile Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly

485 490 495

Asp Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala

500 505 510

Tyr Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg

515 520 525

Val Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val

530 535 540

Phe Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser

545 550 555 560

Leu Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly

565 570 575

Thr Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu

580 585 590

Ser Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr

595 600 605

Ala Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val

610 615 620

Asn Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val

625 630 635 640

Thr Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser

645 650 655

Asp Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys

660 665 670

His Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn

675 680 685

Phe Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr

690 695 700

Asp Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val

705 710 715 720

Thr Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln

725 730 735

Lys Ile Asp Glu Ser Lys Leu Lys Ala Phe Thr Arg Tyr Gln Leu Arg

740 745 750

Gly Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr

755 760 765

Asn Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp

770 775 780

Pro Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg

785 790 795 800

Cys Ala Pro His Leu Glu Trp Asn Pro Asp Leu Asp Cys Ser Cys Arg

805 810 815

Asp Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile

820 825 830

Asp Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile

835 840 845

Phe Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu

850 855 860

Phe Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys

865 870 875 880

Arg Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Glu Trp Glu

885 890 895

Thr Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe

900 905 910

Val Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met

915 920 925

Ile His Ala Ala Asp Lys Arg Val His Ser Ile Arg Glu Ala Tyr Leu

930 935 940

Pro Glu Leu Ser Val Ile Pro Gly Val Asn Ala Ala Ile Phe Glu Glu

945 950 955 960

Leu Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn

965 970 975

Val Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Ser Cys Trp Asn Val

980 985 990

Lys Gly His Val Asp Val Glu Glu Gln Asn Asn Gln Arg Ser Val Leu

995 1000 1005

Val Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val

1010 1015 1020

Cys Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu

1025 1030 1035

Gly Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn

1040 1045 1050

Thr Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Ile Tyr

1055 1060 1065

Pro Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Val Asn Gln Glu

1070 1075 1080

Glu Tyr Gly Gly Ala Tyr Thr Ser Arg Asn Arg Gly Tyr Asn Glu

1085 1090 1095

Ala Pro Ser Val Pro Ala Asp Tyr Ala Ser Val Tyr Glu Glu Lys

1100 1105 1110

Ser Tyr Thr Asp Gly Arg Arg Glu Asn Pro Cys Glu Phe Asn Arg

1115 1120 1125

Gly Tyr Arg Asp Tyr Thr Pro Leu Pro Val Gly Tyr Val Thr Lys

1130 1135 1140

Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp Ile Glu Ile

1145 1150 1155

Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val Glu Leu Leu

1160 1165 1170

Leu Met Glu Glu

1175

<210> 5

<211> 3534

<212> DNA

<213> Artificial sequence

<220>

<223> Fully synthetic sequence encoding BCW 003

<220>

<221> CDS

<222> (1)..(3531)

<400> 5

atg gat aac aat ccg aac atc aat gaa tgc att cct tat aat tgt tta 48

Met Asp Asn Asn Pro Asn Ile Asn Glu Cys Ile Pro Tyr Asn Cys Leu

1 5 10 15

agt aac cct gaa gta gaa gta tta ggt gga gaa aga ata gaa act ggt 96

Ser Asn Pro Glu Val Glu Val Leu Gly Gly Glu Arg Ile Glu Thr Gly

20 25 30

tac acc cca atc gat att tcc ttg tcg cta acg caa ttt ctt ttg agt 144

Tyr Thr Pro Ile Asp Ile Ser Leu Ser Leu Thr Gln Phe Leu Leu Ser

35 40 45

gaa ttt gtt ccc ggt gct gga ttt gtg tta gga cta gtt gat ata ata 192

Glu Phe Val Pro Gly Ala Gly Phe Val Leu Gly Leu Val Asp Ile Ile

50 55 60

tgg gga att ttt ggt ccc tct caa tgg gac gca ttt ctt gta caa att 240

Trp Gly Ile Phe Gly Pro Ser Gln Trp Asp Ala Phe Leu Val Gln Ile

65 70 75 80

gaa cag tta att aac caa aga ata gaa gaa ttc gct agg aac caa gcc 288

Glu Gln Leu Ile Asn Gln Arg Ile Glu Glu Phe Ala Arg Asn Gln Ala

85 90 95

att tct aga tta gaa gga cta agc aat ctt tat caa att tac gca gaa 336

Ile Ser Arg Leu Glu Gly Leu Ser Asn Leu Tyr Gln Ile Tyr Ala Glu

100 105 110

tct ttt aga gag tgg gaa gca gat cct act aat cca gca tta aga gaa 384

Ser Phe Arg Glu Trp Glu Ala Asp Pro Thr Asn Pro Ala Leu Arg Glu

115 120 125

gag atg cgt att caa ttc aat gac atg aac agt gcc ctt aca acc gct 432

Glu Met Arg Ile Gln Phe Asn Asp Met Asn Ser Ala Leu Thr Thr Ala

130 135 140

att cct ctt ttg gca gtt caa aat tat caa gtt cct ctt tta tca gta 480

Ile Pro Leu Leu Ala Val Gln Asn Tyr Gln Val Pro Leu Leu Ser Val

145 150 155 160

tat gtt caa gct gca aat tta cat tta tca gtt ttg aga gat gtt tca 528

Tyr Val Gln Ala Ala Asn Leu His Leu Ser Val Leu Arg Asp Val Ser

165 170 175

gtg ttt gga caa agg tgg gga ttt gat gcc gcg act atc aat agt cgt 576

Val Phe Gly Gln Arg Trp Gly Phe Asp Ala Ala Thr Ile Asn Ser Arg

180 185 190

tat aat gat tta act agg ctt att ggc aac tat aca gat tat gct gta 624

Tyr Asn Asp Leu Thr Arg Leu Ile Gly Asn Tyr Thr Asp Tyr Ala Val

195 200 205

cgc tgg tac aat acg gga tta gaa cgt gta tgg gga ccg gat tct aga 672

Arg Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg

210 215 220

gat tgg gta agg tat aat caa ttt aga aga gaa tta aca cta act gta 720

Asp Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val

225 230 235 240

tta gat atc gtt gct ctg ttc ccg aat tat gat agt aga aga tat cca 768

Leu Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro

245 250 255

att cga aca gtt tcc caa tta aca aga gaa att tat acg aac cca gta 816

Ile Arg Thr Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val

260 265 270

tta gaa aat ttt gat ggt agt ttt cgt gga atg gct cag aga ata gaa 864

Leu Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu

275 280 285

cag aat att agg caa cca cat ctt atg gat atc ctt aat agt ata acc 912

Gln Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr

290 295 300

att tat act gat gtg cat aga ggc ttt aat tat tgg tca ggg cat caa 960

Ile Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln

305 310 315 320

ata aca gct tct cct gta ggg ttt tca gga cca gaa ttc gca ttc cct 1008

Ile Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro

325 330 335

tta ttt ggg aat gcg gga aat gca gct cca ccc gta ctt gtc tca tta 1056

Leu Phe Gly Asn Ala Gly Asn Ala Ala Pro Val Leu Val Ser Leu

340 345 350

act ggt ttg ggg att ttt aga aca tta tct tca cct tta tat aga aga 1104

Thr Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg

355 360 365

att ata ctt ggt tca ggc cca aat aat cag gaa ctg ttt gtc ctt gat 1152

Ile Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp

370 375 380

gga acg gag ttt tct ttt gcc tcc cta acg acc aac ttg cct tcc act 1200

Gly Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr

385 390 395 400

ata tat aga caa agg ggt aca gtc gat tca cta gat gta ata ccg cca 1248

Ile Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro

405 410 415

cag gat aat agt gta cca cct cgt gcg gga ttt agc cat cga ttg agt 1296

Gln Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser

420 425 430

cat gtt aca atg ctg agc caa gca gct gga gca gtt tac acc ttg aga 1344

His Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg

435 440 445

gct cca acg ttt tct tgg cat cgc agt gct acg aca act aat ata 1392

Ala Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile

450 455 460

att gca gcg gat agt att act caa att cct gct gtt aaa gga cgt tct 1440

Ile Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser

465 470 475 480

att att aat aat ggc acg gta att tca gga cca ggg ttt acc gga ggc 1488

Ile Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly

485 490 495

gat ttg gtt aga tta tac aat gct gat ttt aat att aat aat aga gca 1536

Asp Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala

500 505 510

tac ctt gaa gtt ccg ata ttc ttc caa tca ccc tct aca aat tat cgt 1584

Tyr Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg

515 520 525

gtt cgt gtt cgt tat gct tct aca tct tca ctc cct gta gat gta gtt 1632

Val Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val

530 535 540

ttc gga aat att agt cat cct act aca ttc cca gcc act gcc aga tca 1680

Phe Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser

545 550 555 560

tta gat aat cta caa tcc aat gat ttt gga tat att gat att gct gga 1728

Leu Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly

565 570 575

act ttc tta cct tca cta ggg cct agt ata ggt atc aga cct atg tta 1776

Thr Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu

580 585 590

tct act att aat ttg ata gta gat aga ttt gaa ttt att cca gta act 1824

Ser Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr

595 600 605

gca acc ttt gaa gca gaa tcg gat tta gaa aga gca caa aag gcg gtg 1872

Ala Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val

610 615 620

aat gcg ctg ttt act tct aca aac caa cta ggg ata aaa aca gat gtg 1920

Asn Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val

625 630 635 640

acg gat tat cat att gat caa gtg tcc aat tta gtg gag tgt tta tcg 1968

Thr Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser

645 650 655

gat gaa ttt tat ctg gat gaa aag cga gaa ttg tcc gag aaa gtc aaa 2016

Asp Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys

660 665 670

cat gcg aag cga ctc agt gat gag cga aat tta ctt caa gat cca aac 2064

His Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn

675 680 685

ttc agg ggc atc aat aga caa cca gat cgt ggc tgg aga gga agt acg 2112

Phe Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr

690 695 700

gat att acc atc caa gga gga gat gac gta ttc aaa gag aat tac gtc 2160

Asp Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val

705 710 715 720

aca cta cca ggt acc ttt gat gag tgc tat cca aca tat ttg tat caa 2208

Thr Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln

725 730 735

aaa atc gat gaa tca aaa tta aaa gcc ttt acc cgt tat caa tta aga 2256

Lys Ile Asp Glu Ser Lys Leu Lys Ala Phe Thr Arg Tyr Gln Leu Arg

740 745 750

ggg tat atc gaa gat agt caa gac tta gaa atc tat tta att cgc tac 2304

Gly Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr

755 760 765

aat gca aaa cat gaa aca gta aat gtg cca ggt acg ggt tcc tta tgg 2352

Asn Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp

770 775 780

ccg ctt tca gcc caa agt cca atc gga aag tgt gga gag ccg aat cga 2400

Pro Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg

785 790 795 800

tgc gcg cca cac ctt gaa tgg aat cct gac tta gat tgt tcg tgt agg 2448

Cys Ala Pro His Leu Glu Trp Asn Pro Asp Leu Asp Cys Ser Cys Arg

805 810 815

gat gga gaa aag tgt gcc cat cat tcg cat cat ttc tcc tta gac att 2496

Asp Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile

820 825 830

gat gta gga tgt aca gac tta aat gag gac cta ggt gta tgg gtg atc 2544

Asp Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile

835 840 845

ttt aag att aag acg caa gat ggg cac gca aga cta ggg aat cta gag 2592

Phe Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu

850 855 860

ttt ctc gaa gag aaa cca tta gta gga gaa gca cta gct cgt gtg aaa 2640

Phe Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys

865 870 875 880

aga gcg gag aaa aaa tgg aga gac aaa cgt gaa aaa ttg gaa tgg gaa 2688

Arg Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Glu Trp Glu

885 890 895

aca aat atc gtt tat aaa gag gca aaa gaa tct gta gat gct tta ttt 2736

Thr Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe

900 905 910

gta aac tct caa tat gat caa tta caa gcg gat acg aat att gcc atg 2784

Val Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met

915 920 925

att cat gcg gca gat aaa cgt gtt cat agc att cga gaa gct tat ctg 2832

Ile His Ala Ala Asp Lys Arg Val His Ser Ile Arg Glu Ala Tyr Leu

930 935 940

cct gag ctg tct gtg att ccg ggt gtc aat gcg gct att ttt gaa gaa 2880

Pro Glu Leu Ser Val Ile Pro Gly Val Asn Ala Ala Ile Phe Glu Glu

945 950 955 960

tta gaa ggg cgt att ttc act gca ttc tcc cta tat gat gcg aga aat 2928

Leu Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn

965 970 975

gtc att aaa aat ggt gat ttt aat aat ggc tta tcc tgc tgg aac gtg 2976

Val Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Ser Cys Trp Asn Val

980 985 990

aaa ggg cat gta gt gta gaa gaa caa aac aac caa cgt tcg gtc ctt 3024

Lys Gly His Val Asp Val Glu Glu Gln Asn Asn Gln Arg Ser Val Leu

995 1000 1005

gtt gtt ccg gaa tgg gaa gca gaa gtg tca caa gaa gtt cgt gtc 3069

Val Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val

1010 1015 1020

tgt ccg ggt cgt ggc tat atc ctt cgt gtc aca gcg tac aag gag 3114

Cys Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu

1025 1030 1035

gga tat gga gaa ggt tgc gta acc att cat gag atc gag aac aat 3159

Gly Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn

1040 1045 1050

aca gac gaa ctg aag ttt agc aac tgc gta gaa gag gaa atc tat 3204

Thr Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Ile Tyr

1055 1060 1065

cca aat aac acg gta acg tgt aat gat tat act gta aat caa gaa 3249

Pro Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Val Asn Gln Glu

1070 1075 1080

gaa tac gga ggt gcg tac act tct cgt aat cga gga tat aac gaa 3294

Glu Tyr Gly Gly Ala Tyr Thr Ser Arg Asn Arg Gly Tyr Asn Glu

1085 1090 1095

gct cct tcc gta cca gct gat tat gcg tca gtc tat gaa gaa aaa 3339

Ala Pro Ser Val Pro Ala Asp Tyr Ala Ser Val Tyr Glu Glu Lys

1100 1105 1110

tcg tat aca gat gga cga aga gag aat cct tgt gaa ttt aac aga 3384

Ser Tyr Thr Asp Gly Arg Arg Glu Asn Pro Cys Glu Phe Asn Arg

1115 1120 1125

ggg tat agg gat tac acg cca cta cca gtt ggt tat gtg aca aaa 3429

Gly Tyr Arg Asp Tyr Thr Pro Leu Pro Val Gly Tyr Val Thr Lys

1130 1135 1140

gaa tta gaa tac ttc cca gaa acc gat aag gta tgg att gag att 3474

Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp Ile Glu Ile

1145 1150 1155

gga gaa acg gaa gga aca ttt atc gtg gac agc gtg gaa tta ctc 3519

Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val Glu Leu Leu

1160 1165 1170

ctt atg gag gaa tag 3534

Leu Met Glu Glu

1175

<210> 6

<211> 1177

<212> PROTEIN

<213> Artificial sequence

<220>

<223> Synthetic construction

<400> 6

Met Asp Asn Asn Pro Asn Ile Asn Glu Cys Ile Pro Tyr Asn Cys Leu

1 5 10 15

Ser Asn Pro Glu Val Glu Val Leu Gly Gly Glu Arg Ile Glu Thr Gly

20 25 30

Tyr Thr Pro Ile Asp Ile Ser Leu Ser Leu Thr Gln Phe Leu Leu Ser

35 40 45

Glu Phe Val Pro Gly Ala Gly Phe Val Leu Gly Leu Val Asp Ile Ile

50 55 60

Trp Gly Ile Phe Gly Pro Ser Gln Trp Asp Ala Phe Leu Val Gln Ile

65 70 75 80

Glu Gln Leu Ile Asn Gln Arg Ile Glu Glu Phe Ala Arg Asn Gln Ala

85 90 95

Ile Ser Arg Leu Glu Gly Leu Ser Asn Leu Tyr Gln Ile Tyr Ala Glu

100 105 110

Ser Phe Arg Glu Trp Glu Ala Asp Pro Thr Asn Pro Ala Leu Arg Glu

115 120 125

Glu Met Arg Ile Gln Phe Asn Asp Met Asn Ser Ala Leu Thr Thr Ala

130 135 140

Ile Pro Leu Leu Ala Val Gln Asn Tyr Gln Val Pro Leu Leu Ser Val

145 150 155 160

Tyr Val Gln Ala Ala Asn Leu His Leu Ser Val Leu Arg Asp Val Ser

165 170 175

Val Phe Gly Gln Arg Trp Gly Phe Asp Ala Ala Thr Ile Asn Ser Arg

180 185 190

Tyr Asn Asp Leu Thr Arg Leu Ile Gly Asn Tyr Thr Asp Tyr Ala Val

195 200 205

Arg Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg

210 215 220

Asp Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val

225 230 235 240

Leu Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro

245 250 255

Ile Arg Thr Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val

260 265 270

Leu Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu

275 280 285

Gln Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr

290 295 300

Ile Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln

305 310 315 320

Ile Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro

325 330 335

Leu Phe Gly Asn Ala Gly Asn Ala Ala Pro Val Leu Val Ser Leu

340 345 350

Thr Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg

355 360 365

Ile Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp

370 375 380

Gly Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr

385 390 395 400

Ile Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro

405 410 415

Gln Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser

420 425 430

His Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg

435 440 445

Ala Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile

450 455 460

Ile Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser

465 470 475 480

Ile Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly

485 490 495

Asp Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala

500 505 510

Tyr Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg

515 520 525

Val Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val

530 535 540

Phe Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser

545 550 555 560

Leu Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly

565 570 575

Thr Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu

580 585 590

Ser Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr

595 600 605

Ala Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val

610 615 620

Asn Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val

625 630 635 640

Thr Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser

645 650 655

Asp Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys

660 665 670

His Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn

675 680 685

Phe Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr

690 695 700

Asp Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val

705 710 715 720

Thr Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln

725 730 735

Lys Ile Asp Glu Ser Lys Leu Lys Ala Phe Thr Arg Tyr Gln Leu Arg

740 745 750

Gly Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr

755 760 765

Asn Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp

770 775 780

Pro Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg

785 790 795 800

Cys Ala Pro His Leu Glu Trp Asn Pro Asp Leu Asp Cys Ser Cys Arg

805 810 815

Asp Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile

820 825 830

Asp Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile

835 840 845

Phe Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu

850 855 860

Phe Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys

865 870 875 880

Arg Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Glu Trp Glu

885 890 895

Thr Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe

900 905 910

Val Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met

915 920 925

Ile His Ala Ala Asp Lys Arg Val His Ser Ile Arg Glu Ala Tyr Leu

930 935 940

Pro Glu Leu Ser Val Ile Pro Gly Val Asn Ala Ala Ile Phe Glu Glu

945 950 955 960

Leu Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn

965 970 975

Val Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Ser Cys Trp Asn Val

980 985 990

Lys Gly His Val Asp Val Glu Glu Gln Asn Asn Gln Arg Ser Val Leu

995 1000 1005

Val Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val

1010 1015 1020

Cys Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu

1025 1030 1035

Gly Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn

1040 1045 1050

Thr Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Ile Tyr

1055 1060 1065

Pro Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Val Asn Gln Glu

1070 1075 1080

Glu Tyr Gly Gly Ala Tyr Thr Ser Arg Asn Arg Gly Tyr Asn Glu

1085 1090 1095

Ala Pro Ser Val Pro Ala Asp Tyr Ala Ser Val Tyr Glu Glu Lys

1100 1105 1110

Ser Tyr Thr Asp Gly Arg Arg Glu Asn Pro Cys Glu Phe Asn Arg

1115 1120 1125

Gly Tyr Arg Asp Tyr Thr Pro Leu Pro Val Gly Tyr Val Thr Lys

1130 1135 1140

Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp Ile Glu Ile

1145 1150 1155

Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val Glu Leu Leu

1160 1165 1170

Leu Met Glu Glu

1175

<210> 7

<211> 3543

<212> DNA

<213> Artificial sequence

<220>

<223> Fully synthetic sequence for application

in plants, encoding BCW 001

<220>

<221> CDS

<222> (1)..(3540)

<223> BCW 001

<400> 7

atg gag gag aac aac cag aac cag tgt gtc cca tac aac tgc ctc aac 48

Met Glu Glu Asn Asn Gln Asn Gln Cys Val Pro Tyr Asn Cys Leu Asn

1 5 10 15

aac cca gcc atc gaa atc ctt gag ggc gac cga att tca gtc ggc aac 96

Asn Pro Ala Ile Glu Ile Leu Glu Gly Asp Arg Ile Ser Val Gly Asn

20 25 30

acg ccc atc gac atc tcc ctg agt ctt gtg gaa ctc ctc atc tcg gag 144

Thr Pro Ile Asp Ile Ser Leu Ser Leu Val Glu Leu Leu Ile Ser Glu

35 40 45

ttc gtc cct ggc ggc ggc ata atc acc ggt ctg ctc aac atc gtg tgg 192

Phe Val Pro Gly Gly Gly Ile Ile Thr Gly Leu Leu Asn Ile Val Trp

50 55 60

gga ttc gtg ggc cca tcc cag tgg gat gcg ttc ctg gcc caa gtg gag 240

Gly Phe Val Gly Pro Ser Gln Trp Asp Ala Phe Leu Ala Gln Val Glu

65 70 75 80

cag ctc atc aac cag agg atc tcc gag gcc gtc cgc aat acc gcg atc 288

Gln Leu Ile Asn Gln Arg Ile Ser Glu Ala Val Arg Asn Thr Ala Ile

85 90 95

caa gag ctg gag ggc atg gcc cgc gtg tac cgc acc tac gcc acc gcc 336

Gln Glu Leu Glu Gly Met Ala Arg Val Tyr Arg Thr Tyr Ala Thr Ala

100 105 110

ttt gct gaa tgg gag cgc gac ccg aac aac act gac ctg cgc gag gcc 384

Phe Ala Glu Trp Glu Arg Asp Pro Asn Asn Thr Asp Leu Arg Glu Ala

115 120 125

gtc cga aca cag ttc acg gcg acc gag acc tac atc agc ggc cgg atc 432

Val Arg Thr Gln Phe Thr Ala Thr Glu Thr Tyr Ile Ser Gly Arg Ile

130 135 140

tca gtg ctc aag atc cag aac ttc gag gtg cag ctc cta tcg gtc ttc 480

Ser Val Leu Lys Ile Gln Asn Phe Glu Val Gln Leu Leu Ser Val Phe

145 150 155 160

gcc cag gcc gcc aac ttg cac ctg agc ctc ctg cgg gac gtt gtg ttc 528

Ala Gln Ala Ala Asn Leu His Leu Ser Leu Leu Arg Asp Val Val Phe

165 170 175

ttc ggc cag cgg tgg ggc ttc tct act acg acc gtg aac aac tac tac 576

Phe Gly Gln Arg Trp Gly Phe Ser Thr Thr Thr Val Asn Asn Tyr Tyr

180 185 190

aac gac ctg acg gag gaa atc agc acc tac aca gat tac gca gtt cgt 624

Asn Asp Leu Thr Glu Glu Ile Ser Thr Tyr Thr Asp Tyr Ala Val Arg

195 200 205

tgg tac aac acc ggc ctt gag cgc gtg tgg ggc ccg gac tcc cgc gat 672

Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg Asp

210 215 220

tgg gtc cgc tac aac cag ttc cgc cgc gag ctg acg ctt aca gtg ctg 720

Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val Leu

225 230 235 240

gac atc gtc gca ctc ttt cct aac tac gac tcc agg cgc tat ccc atc 768

Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro Ile

245 250 255

agg aca gtg tca cag ctc acc cgc gag atc tac aca aac ccg gtg ctt 816

Arg Thr Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val Leu

260 265 270

gag aac ttc gac ggc agc ttc cgt ggc atg gcg cag cgc att gaa cag 864

Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu Gln

275 280 285

aac atc cgc cag cg cac ctt atg gac atc ttg aac agt atc act atc 912

Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr Ile

290 295 300

tac acc gac gtc cac aga ggc ttc aac tac tgg agc gga cac cag atc 960

Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln Ile

305 310 315 320

aca gcc agc ccg gta ggc ttc tcg ggt cca gag ttc gcc ttc ccg ctg 1008

Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro Leu

325 330 335

ttt ggg aac gct ggc aat gcc gcg ccg ccc gtg ctg gtc agc ctc act 1056

Phe Gly Asn Ala Gly Asn Ala Ala Pro Pro Val Leu Val Ser Leu Thr

340 345 350

ggt ctc ggc atc ttc cgc aca ctt tcc tcg ccg ctg tac agg agg atc 1104

Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg Ile

355 360 365

atc ctc ggg tcc ggt ccg aac aac cag gag ctg ttc gtg ctc gac ggg 1152

Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp Gly

370 375 380

acc gag ttc agt ttc gcc agc ctc acg acg aac ctc ccg tcc acc atc 1200

Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr Ile

385 390 395 400

tat cgc cag cgc gga acg gtc gat tcc ctg gat gtt atc cca ccg caa 1248

Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro Gln

405 410 415

gac aat tct gtg ccg ccg agg gcc ggg ttc tcc cac cgg ctg tct cac 1296

Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser His

420 425 430

gtg act atg ctt tca cag gcc gcc gga gcc gtg tac acg ctc cgt gcg 1344

Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg Ala

435 440 445

cct act ttc tcc tgg cag cac cgc agc gcg acc acg acc aac atc atc 1392

Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile Ile

450 455 460

gca gca gac tcc atc acc cag atc ccg gcc gtt aag ggc cgc agc atc 1440

Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser Ile

465 470 475 480

atc aac aac gga act gtc atc agc ggt ccg ggc ttc acg ggc ggc gac 1488

Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly Asp

485 490 495

ctg gtc cgg ctc tac aac gca gac ttc aac atc aat aac cgc gct tat 1536

Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala Tyr

500 505 510

ctt gaa gta cct atc ttc ttc cag agc ccg agc act aac tac cgg gtt 1584

Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg Val

515 520 525

cgc gtc cgc tac gcc agc acc tcc agc ctc cct gtg gat gtc gtg ttc 1632

Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val Phe

530 535 540

gga aac ata agc cat ccg acc acg ttc cca gcc acg gct agg agc ctg 1680

Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser Leu

545 550 555 560

gac aac cta cag agc aac gac ttc ggc tac atc gac atc gcg ggc acc 1728

Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly Thr

565 570 575

ttt ctg cca agc ctg ggt ccg tct atc ggc atc cgc ccg atg ctg agc 1776

Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu Ser

580 585 590

act atc aac cta att gtg gac cgg ttc gag ttt atc ccg gtg acg gca 1824

Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr Ala

595 600 605

acg ttc gag gcg gag tct gac ctc gaa agg gca cag aag gcc gtg aac 1872

Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val Asn

610 615 620

gcc ctg ttc acg agc acc aac cag ctt ggc att aag act gat gtc acc 1920

Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val Thr

625 630 635 640

gac tac cac att gac caa gtc agc aac ctg gtg gag tgc ctc tcg gac 1968

Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser Asp

645 650 655

gag ttc tat ctt gat gag aaa cgg gaa cta agc gag aag gtg aag cac 2016

Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys His

660 665 670

gca aag cgc ttg agc gac gag cgg aac tta ctc cag gac cct aac ttc 2064

Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn Phe

675 680 685

cgt ggg att aac cgc cag ccg gat cgc ggg tgg cgc ggc tca acg gac 2112

Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr Asp

690 695 700

atc acc atc cag ggc ggc gat gac gtc ttc aag gag aac tac gtg acc 2160

Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val Thr

705 710 715 720

ctc cct ggc acg ttc gac gag tgc tac ccg acg tac ctt tat cag aag 2208

Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln Lys

725 730 735

att gac gaa agc aag ctg aaa gcc tac acc cgc tac cag ttg cgc ggc 2256

Ile Asp Glu Ser Lys Leu Lys Ala Tyr Thr Arg Tyr Gln Leu Arg Gly

740 745 750

tac atc gag gac tct caa gac ctg gag atc tac ttg att cga tac aac 2304

Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr Asn

755 760 765

gcg aaa cac gag acc gtc aac gtg ccg ggc act ggg agc ctg tgg ccg 2352

Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp Pro

770 775 780

ttg tct gca caa agt ccg atc ggc aag tgc ggc gag cca aac cgg tgc 2400

Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg Cys

785 790 795 800

gct ccg cac ctg gag tgg aac cca gac ctt cat tgc tcc tgt agg gat 2448

Ala Pro His Leu Glu Trp Asn Pro Asp Leu His Cys Ser Cys Arg Asp

805 810 815

ggc gag aag tgc gct cac cac agc cat cac ttc agc ctc gac att gac 2496

Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile Asp

820 825 830

gtc ggc tgt acc gac ctt aat gag gat ctg ggt gtg tgg gtg atc ttc 2544

Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile Phe

835 840 845

aag atc aag acc cag gac ggt cac gcc cgg ttg ggc aat ctg gag ttc 2592

Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu Phe

850 855 860

ctg gag gag aag ccg ctg gtt ggc gag gct ctc gcg cgg gtc aag cgg 2640

Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys Arg

865 870 875 880

gcg gag aag aag tgg cgg gac aaa cgc gag aag ctc cag tta gag acg 2688

Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Gln Leu Glu Thr

885 890 895

aac atc gtg tac aag gag gcg aag gaa tcc gtg gac gca cta ttc gtg 2736

Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe Val

900 905 910

aac agc cag tac gac caa ctc cag gcc gac acc aac atc gcc atg att 2784

Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met Ile

915 920 925

cac gca gcc gac aag agg gtg cac cgc atc cgc gaa gcc tac ctt ccc 2832

His Ala Ala Asp Lys Arg Val His Arg Ile Arg Glu Ala Tyr Leu Pro

930 935 940

gaa ctt tcg gtc atc cca ggc gtc aac gct gac atc tcg gag gaa ttg 2880

Glu Leu Ser Val Ile Pro Gly Val Asn Ala Asp Ile Ser Glu Glu Leu

945 950 955 960

gag ggc aga atc ttc acg gcc ttc tct ttg tac gat gcc agg aac gtc 2928

Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn Val

965 970 975

atc aag aac ggc gac ttc aac aac ggc ctg ctg tgc tgg aac gtg aag 2976

Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Leu Cys Trp Asn Val Lys

980 985 990

ggc cac gtg gac gtc gag gag cag aac aac cac aga tca gtc ctg gtg 3024

Gly His Val Asp Val Glu Glu Gln Asn Asn His Arg Ser Val Leu Val

995 1000 1005

gtg ccc gag tgg gaa gcc gaa gtc tca caa gaa gtc cgg gtg tgc 3069

Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val Cys

1010 1015 1020

cct gga cgc ggg tac att ctc cgc gtg acc gcc tac aag gag ggc 3114

Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu Gly

1025 1030 1035

tac ggt gag ggc tgc gtg acc atc cac gag atc gag aac aac acc 3159

Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn Thr

1040 1045 1050

gac gag ctg aaa ttc agt aac tgt gtt gag gag gag gtg tac ccg 3204

Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Val Tyr Pro

1055 1060 1065

aac aac acc gtc acc tgc aac gac tac act gcg aac cag gag gaa 3249

Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Ala Asn Gln Glu Glu

1070 1075 1080

tac gag ggc acg tac acg agc cgc aat cgc ggg tac gac gag gcg 3294

Tyr Glu Gly Thr Tyr Thr Ser Arg Asn Arg Gly Tyr Asp Glu Ala

1085 1090 1095

tac gag agc aac tcc agc gtc ccg gcc gag tac gcc tcc gtg tac 3339

Tyr Glu Ser Asn Ser Ser Val Pro Ala Glu Tyr Ala Ser Val Tyr

1100 1105 1110

gag gag aag gtt tac acc gac ggg agg cgt ggc aac ccg tgc gag 3384

Glu Glu Lys Val Tyr Thr Asp Gly Arg Arg Gly Asn Pro Cys Glu

1115 1120 1125

agc aac aga ggc tac ggc gat tac act ccg ctt ccc gct ggc tac 3429

Ser Asn Arg Gly Tyr Gly Asp Tyr Thr Pro Leu Pro Ala Gly Tyr

1130 1135 1140

gtg acg aaa gag ctg gag tac ttc cca gag acc gac aag gtg tgg 3474

Val Thr Lys Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp

1145 1150 1155

atc gag atc gga gaa acg gag ggc acg ttc ata gtg gac tcc gtt 3519

Ile Glu Ile Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val

1160 1165 1170

gag ctg ctg ctc atg gag gag tag 3543

Glu Leu Leu Leu Met Glu Glu

1175 1180

<210> 8

<211> 1180

<212> PROTEIN

<213> Artificial sequence

<220>

<223> Synthetic construction

<400> 8

Met Glu Glu Asn Asn Gln Asn Gln Cys Val Pro Tyr Asn Cys Leu Asn

1 5 10 15

Asn Pro Ala Ile Glu Ile Leu Glu Gly Asp Arg Ile Ser Val Gly Asn

20 25 30

Thr Pro Ile Asp Ile Ser Leu Ser Leu Val Glu Leu Leu Ile Ser Glu

35 40 45

Phe Val Pro Gly Gly Gly Ile Ile Thr Gly Leu Leu Asn Ile Val Trp

50 55 60

Gly Phe Val Gly Pro Ser Gln Trp Asp Ala Phe Leu Ala Gln Val Glu

65 70 75 80

Gln Leu Ile Asn Gln Arg Ile Ser Glu Ala Val Arg Asn Thr Ala Ile

85 90 95

Gln Glu Leu Glu Gly Met Ala Arg Val Tyr Arg Thr Tyr Ala Thr Ala

100 105 110

Phe Ala Glu Trp Glu Arg Asp Pro Asn Asn Thr Asp Leu Arg Glu Ala

115 120 125

Val Arg Thr Gln Phe Thr Ala Thr Glu Thr Tyr Ile Ser Gly Arg Ile

130 135 140

Ser Val Leu Lys Ile Gln Asn Phe Glu Val Gln Leu Leu Ser Val Phe

145 150 155 160

Ala Gln Ala Ala Asn Leu His Leu Ser Leu Leu Arg Asp Val Val Phe

165 170 175

Phe Gly Gln Arg Trp Gly Phe Ser Thr Thr Thr Val Asn Asn Tyr Tyr

180 185 190

Asn Asp Leu Thr Glu Glu Ile Ser Thr Tyr Thr Asp Tyr Ala Val Arg

195 200 205

Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg Asp

210 215 220

Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val Leu

225 230 235 240

Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro Ile

245 250 255

Arg Thr Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val Leu

260 265 270

Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu Gln

275 280 285

Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr Ile

290 295 300

Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln Ile

305 310 315 320

Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro Leu

325 330 335

Phe Gly Asn Ala Gly Asn Ala Ala Pro Pro Val Leu Val Ser Leu Thr

340 345 350

Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg Ile

355 360 365

Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp Gly

370 375 380

Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr Ile

385 390 395 400

Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro Gln

405 410 415

Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser His

420 425 430

Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg Ala

435 440 445

Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile Ile

450 455 460

Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser Ile

465 470 475 480

Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly Asp

485 490 495

Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala Tyr

500 505 510

Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg Val

515 520 525

Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val Phe

530 535 540

Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser Leu

545 550 555 560

Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly Thr

565 570 575

Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu Ser

580 585 590

Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr Ala

595 600 605

Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val Asn

610 615 620

Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val Thr

625 630 635 640

Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser Asp

645 650 655

Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys His

660 665 670

Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn Phe

675 680 685

Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr Asp

690 695 700

Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val Thr

705 710 715 720

Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln Lys

725 730 735

Ile Asp Glu Ser Lys Leu Lys Ala Tyr Thr Arg Tyr Gln Leu Arg Gly

740 745 750

Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr Asn

755 760 765

Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp Pro

770 775 780

Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg Cys

785 790 795 800

Ala Pro His Leu Glu Trp Asn Pro Asp Leu His Cys Ser Cys Arg Asp

805 810 815

Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile Asp

820 825 830

Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile Phe

835 840 845

Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu Phe

850 855 860

Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys Arg

865 870 875 880

Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Gln Leu Glu Thr

885 890 895

Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe Val

900 905 910

Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met Ile

915 920 925

His Ala Ala Asp Lys Arg Val His Arg Ile Arg Glu Ala Tyr Leu Pro

930 935 940

Glu Leu Ser Val Ile Pro Gly Val Asn Ala Asp Ile Ser Glu Glu Leu

945 950 955 960

Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn Val

965 970 975

Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Leu Cys Trp Asn Val Lys

980 985 990

Gly His Val Asp Val Glu Glu Gln Asn Asn His Arg Ser Val Leu Val

995 1000 1005

Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val Cys

1010 1015 1020

Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu Gly

1025 1030 1035

Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn Thr

1040 1045 1050

Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Val Tyr Pro

1055 1060 1065

Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Ala Asn Gln Glu Glu

1070 1075 1080

Tyr Glu Gly Thr Tyr Thr Ser Arg Asn Arg Gly Tyr Asp Glu Ala

1085 1090 1095

Tyr Glu Ser Asn Ser Ser Val Pro Ala Glu Tyr Ala Ser Val Tyr

1100 1105 1110

Glu Glu Lys Val Tyr Thr Asp Gly Arg Arg Gly Asn Pro Cys Glu

1115 1120 1125

Ser Asn Arg Gly Tyr Gly Asp Tyr Thr Pro Leu Pro Ala Gly Tyr

1130 1135 1140

Val Thr Lys Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp

1145 1150 1155

Ile Glu Ile Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val

1160 1165 1170

Glu Leu Leu Leu Met Glu Glu

1175 1180

<210> 9

<211> 3534

<212> DNA

<213> Artificial sequence

<220>

<223> Fully synthetic sequence encoding

CW 002 for plant applications

<220>

<221> CDS

<222> (1)..(3531)

<223> BCW 002

<400> 9

atg gac aac aac ccg aac atc aac gag tgc atc ccc tac aac tgc ctc 48

Met Asp Asn Asn Pro Asn Ile Asn Glu Cys Ile Pro Tyr Asn Cys Leu

1 5 10 15

tcc aac ccg gag gtc gag gtg ctg ggc ggc gaa agg atc gag acc ggc 96

Ser Asn Pro Glu Val Glu Val Leu Gly Gly Glu Arg Ile Glu Thr Gly

20 25 30

tac act ccc atc gac atc agc ctc agc ctg acc cag ttc ctg ctc tct 144

Tyr Thr Pro Ile Asp Ile Ser Leu Ser Leu Thr Gln Phe Leu Leu Ser

35 40 45

gag ttc gtg ccc ggc gcg ggg ttc gtt ctc ggc ctg gtc gac atc atc 192

Glu Phe Val Pro Gly Ala Gly Phe Val Leu Gly Leu Val Asp Ile Ile

50 55 60

tgg ggc atc ttc ggt ccg agc cag tgg gac gcc ttt ctc gtt cag att 240

Trp Gly Ile Phe Gly Pro Ser Gln Trp Asp Ala Phe Leu Val Gln Ile

65 70 75 80

gag cag ctg atc aac cag cgc atc gag gag ttc gcc cgc aac cag gcg 288

Glu Gln Leu Ile Asn Gln Arg Ile Glu Glu Phe Ala Arg Asn Gln Ala

85 90 95

atc tcc cgg ctg gag ggc ctc tcc aac ctg tac caa atc tac gcc gag 336

Ile Ser Arg Leu Glu Gly Leu Ser Asn Leu Tyr Gln Ile Tyr Ala Glu

100 105 110

agc ttc cgg gag tgg gaa gcc gat ccg acc aac ccc gct ctc agg gag 384

Ser Phe Arg Glu Trp Glu Ala Asp Pro Thr Asn Pro Ala Leu Arg Glu

115 120 125

gag atg cgg att cag ttc aac gac atg aac tcc gct ctc acg act gcc 432

Glu Met Arg Ile Gln Phe Asn Asp Met Asn Ser Ala Leu Thr Thr Ala

130 135 140

atc cca ctc ctc gct gtg cag aac tac caa gtg ccg ctc ctg tcc gtg 480

Ile Pro Leu Leu Ala Val Gln Asn Tyr Gln Val Pro Leu Leu Ser Val

145 150 155 160

tac gtg cag gcc gcc aat ctg cac ctc tcc gtc ctc cgg gac gtt agc 528

Tyr Val Gln Ala Ala Asn Leu His Leu Ser Val Leu Arg Asp Val Ser

165 170 175

gtg ttc ggg cag cgc tgg ggc ttc gac gcc gct acc atc aac tcc cgt 576

Val Phe Gly Gln Arg Trp Gly Phe Asp Ala Ala Thr Ile Asn Ser Arg

180 185 190

tac aac gat ctc act cgc ctc atc ggc aac tac acc gac tat gcc gtg 624

Tyr Asn Asp Leu Thr Arg Leu Ile Gly Asn Tyr Thr Asp Tyr Ala Val

195 200 205

cgc tgg tac aac act ggt ctt gag aga gtc tgg ggc ccg gac agc cgc 672

Arg Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg

210 215 220

gac tgg gtg cgc tac aac cag ttc cgg cgc gag ctg acc ctc acc gtg 720

Asp Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val

225 230 235 240

ctc gac atc gta gcc ctc ttt ccc aac tac gac tcc cgg cgc tac ccg 768

Leu Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro

245 250 255

att cgc atc gtc agc cag ctc acc agg gag atc tac acc aac cct gtg 816

Ile Arg Ile Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val

260 265 270

ctg gag aac ttc gac ggc tcc ttt cgc ggg atg gcc caa cgc ata gag 864

Leu Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu

275 280 285

cag aac atc cgc caa cct cat ctg atg gac atc ctt aat tct atc acc 912

Gln Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr

290 295 300

atc tac act gac gtt cat cgc ggg ttt aac tac tgg tcg ggc cac caa 960

Ile Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln

305 310 315 320

atc act gcg tcg ccc gtt ggt ttc tcc ggc ccg gag ttc gcg ttc cct 1008

Ile Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro

325 330 335

ctg ttc gga aac gcg ggc aat gcc gct cca ccc gta ttg gtg agc ctg 1056

Leu Phe Gly Asn Ala Gly Asn Ala Ala Pro Val Leu Val Ser Leu

340 345 350

acc ggc ctc ggc atc ttc cgt aca ctg tct agc cct ctg tac aga agg 1104

Thr Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg

355 360 365

atc att ctt ggc agc ggt ccc aat aac cag gaa ctc ttc gtg ttg gac 1152

Ile Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp

370 375 380

ggc acc gag ttc agc ttc gcc agt ctt acg acc aat ttg ccc tcc aca 1200

Gly Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr

385 390 395 400

atc tat cgc cag cgc ggt act gtg gac tcc ctt gat gtg ata cca cct 1248

Ile Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro

405 410 415

cag gac aac tct gtc cca cct cgc gcc ggt ttc tcc cac cgc ctc agc 1296

Gln Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser

420 425 430

cac gtc act atg ctg agt cag gct gcg gga gcc gtg tac acc ctt cgg 1344

His Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg

435 440 445

gct ccg acg ttt agc tgg cag cac agg agc gcg act acc acg aac atc 1392

Ala Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile

450 455 460

att gcg gct gac tcc atc act caa atc cct gcc gtt aag ggt cgc tcc 1440

Ile Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser

465 470 475 480

atc atc aac aat ggg aca gtg atc tcg gga ccg ggc ttc acc ggc ggt 1488

Ile Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly

485 490 495

gac ctg gtg agg ctg tac aac gcg gac ttc aac atc aac aac agg gcg 1536

Asp Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala

500 505 510

tac ctc gaa gtc ccg atc ttc ttc cag tcg ccc agc acg aac tat cgt 1584

Tyr Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg

515 520 525

gtc agg gtc cgg tac gcc tca acc tca tcc ctc ccg gtc gat gtg gtc 1632

Val Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val

530 535 540

ttc ggc aac atc agc cac ccg acc acg ttt ccg gct acc gcc cga tcc 1680

Phe Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser

545 550 555 560

ctg gac aat ctg caa agc aac gat ttc ggc tac att gac att gcc ggg 1728

Leu Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly

565 570 575

acg ttc ctc ccg agc ctc ggc cca tcc atc ggc atc cgg ccc atg ctc 1776

Thr Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu

580 585 590

tcc acc atc aac ctg atc gtg gat cgg ttt gag ttc atc cca gtg aca 1824

Ser Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr

595 600 605

gcc act ttc gag gct gag tcc gac cta gag cgt gct cag aag gca gtc 1872

Ala Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val

610 615 620

aat gct ctg ttt acc tcc acc aat cag ctc ggc att aag acc gat gtg 1920

Asn Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val

625 630 635 640

acc gat tac cac att gac caa gtc tca aac ctc gtt gag tgc ctc tcg 1968

Thr Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser

645 650 655

gat gag ttc tac ctt gat gag aag agg gag ctt tca gag aaa gtt aag 2016

Asp Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys

660 665 670

cac gct aag aga ctc tcg gac gaa cgc aat ctg ttg caa gat ccc aac 2064

His Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn

675 680 685

ttc aga ggg atc aac cgt cag cca gac cgg gga tgg cgc ggg tcc acg 2112

Phe Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr

690 695 700

gac atc act atc cag ggc ggt gat gac gtc ttc aag gag aac tac gtg 2160

Asp Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val

705 710 715 720

acc ctg ccg ggc acc ttt gac gaa tgc tac ccc act tac ctc tac cag 2208

Thr Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln

725 730 735

aag att gac gag tcc aag ctc aag gcg ttc aca cgc tac cag ctc agg 2256

Lys Ile Asp Glu Ser Lys Leu Lys Ala Phe Thr Arg Tyr Gln Leu Arg

740 745 750

ggt tac atc gag gac tcc caa gac ctg gaa atc tac ctg atc cgc tac 2304

Gly Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr

755 760 765

aac gct aag cac gag act gtc aac gtg ccc ggc acc ggc agc ctg tgg 2352

Asn Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp

770 775 780

ccc ttg tcc gct cag agc cca atc ggc aag tgc ggc gag ccc aac cgc 2400

Pro Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg

785 790 795 800

tgc gcg ccc cac ctg gaa tgg aac ccc gac ctc gac tgt agc tgc cgc 2448

Cys Ala Pro His Leu Glu Trp Asn Pro Asp Leu Asp Cys Ser Cys Arg

805 810 815

gac gga gag aag tgc gcg cat cac tcc cac cac ttc agc ctc gac atc 2496

Asp Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile

820 825 830

gac gtc ggt tgc acc gac ctt aac gag gat ctg ggc gtt tgg gtg atc 2544

Asp Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile

835 840 845

ttc aag atc aag act cag gac ggc cac gcc cgc ctg gga aac ctg gag 2592

Phe Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu

850 855 860

ttc ctg gag gag aag ccc ctc gtt ggc gag gcc ctg gcc cgc gtc aag 2640

Phe Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys

865 870 875 880

agg gcc gag aag aaa tgg cgc gac aag cgc gag aag ctg gag tgg gag 2688

Arg Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Glu Trp Glu

885 890 895

acc aac atc gtg tac aag gaa gcg aag gag tca gtt gac gcc ctg ttc 2736

Thr Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe

900 905 910

gtc aac agc cag tac gac cag ctc cag gca gac aca aac atc gct atg 2784

Val Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met

915 920 925

atc cat gcg gcc gac aag cgc gtc cac tcc atc cgc gag gcg tac ctg 2832

Ile His Ala Ala Asp Lys Arg Val His Ser Ile Arg Glu Ala Tyr Leu

930 935 940

ccc gag ctg tcc gtc atc ccc ggc gtc aac gcc gcg atc ttt gag gag 2880

Pro Glu Leu Ser Val Ile Pro Gly Val Asn Ala Ala Ile Phe Glu Glu

945 950 955 960

ctg gag ggc cgc atc ttc acc gcc ttc tcc ctc tac gac gca cgc aac 2928

Leu Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn

965 970 975

gtt atc aag aat ggc gac ttc aac aac ggg ctg tcc tgc tgg aat gtc 2976

Val Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Ser Cys Trp Asn Val

980 985 990

aag ggc cac gtg gac gtc gag gag cag aac aac cag cgc tca gtc ctg 3024

Lys Gly His Val Asp Val Glu Glu Gln Asn Asn Gln Arg Ser Val Leu

995 1000 1005

gtc gtc ccg gag tgg gag gcc gaa gtc agc cag gaa gtc cgc gtc 3069

Val Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val

1010 1015 1020

tgc cct gga cgc ggg tac atc ctg cgc gtc act gcc tac aag gaa 3114

Cys Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu

1025 1030 1035

ggc tac gga gag ggc tgc gtc acc atc cat gag atc gaa aac aac 3159

Gly Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn

1040 1045 1050

acg gat gag ctt aag ttc agc aac tgt gtt gaa gag gaa atc tac 3204

Thr Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Ile Tyr

1055 1060 1065

ccg aac aac acg gtc acc tgc aat gat tac acc gtc aac cag gag 3249

Pro Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Val Asn Gln Glu

1070 1075 1080

gaa tac ggt gga gct tac acc tcc cgc aac agg ggc tac aac gag 3294

Glu Tyr Gly Gly Ala Tyr Thr Ser Arg Asn Arg Gly Tyr Asn Glu

1085 1090 1095

gca ccc tct gtc ccg gcc gac tac gct tca gtc tac gaa gag aag 3339

Ala Pro Ser Val Pro Ala Asp Tyr Ala Ser Val Tyr Glu Glu Lys

1100 1105 1110

tcg tac acc gac gga cgc aga gag aac ccg tgt gag ttc aac cgc 3384

Ser Tyr Thr Asp Gly Arg Arg Glu Asn Pro Cys Glu Phe Asn Arg

1115 1120 1125

ggc tac cgc gat tac acc ccg ctg cct gtc ggg tac gtc acc aaa 3429

Gly Tyr Arg Asp Tyr Thr Pro Leu Pro Val Gly Tyr Val Thr Lys

1130 1135 1140

gag ctg gaa tac ttc cca gag acc gac aaa gtc tgg att gag atc 3474

Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp Ile Glu Ile

1145 1150 1155

ggc gag acc gag ggc acg ttc atc gtg gac tcc gtc gaa ctc ctt 3519

Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val Glu Leu Leu

1160 1165 1170

ctg atg gaa gag tga 3534

Leu Met Glu Glu

1175

<210> 10

<211> 1177

<212> PROTEIN

<213> Artificial sequence

<220>

<223> Synthetic construction

<400> 10

Met Asp Asn Asn Pro Asn Ile Asn Glu Cys Ile Pro Tyr Asn Cys Leu

1 5 10 15

Ser Asn Pro Glu Val Glu Val Leu Gly Gly Glu Arg Ile Glu Thr Gly

20 25 30

Tyr Thr Pro Ile Asp Ile Ser Leu Ser Leu Thr Gln Phe Leu Leu Ser

35 40 45

Glu Phe Val Pro Gly Ala Gly Phe Val Leu Gly Leu Val Asp Ile Ile

50 55 60

Trp Gly Ile Phe Gly Pro Ser Gln Trp Asp Ala Phe Leu Val Gln Ile

65 70 75 80

Glu Gln Leu Ile Asn Gln Arg Ile Glu Glu Phe Ala Arg Asn Gln Ala

85 90 95

Ile Ser Arg Leu Glu Gly Leu Ser Asn Leu Tyr Gln Ile Tyr Ala Glu

100 105 110

Ser Phe Arg Glu Trp Glu Ala Asp Pro Thr Asn Pro Ala Leu Arg Glu

115 120 125

Glu Met Arg Ile Gln Phe Asn Asp Met Asn Ser Ala Leu Thr Thr Ala

130 135 140

Ile Pro Leu Leu Ala Val Gln Asn Tyr Gln Val Pro Leu Leu Ser Val

145 150 155 160

Tyr Val Gln Ala Ala Asn Leu His Leu Ser Val Leu Arg Asp Val Ser

165 170 175

Val Phe Gly Gln Arg Trp Gly Phe Asp Ala Ala Thr Ile Asn Ser Arg

180 185 190

Tyr Asn Asp Leu Thr Arg Leu Ile Gly Asn Tyr Thr Asp Tyr Ala Val

195 200 205

Arg Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg

210 215 220

Asp Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val

225 230 235 240

Leu Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro

245 250 255

Ile Arg Ile Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val

260 265 270

Leu Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu

275 280 285

Gln Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr

290 295 300

Ile Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln

305 310 315 320

Ile Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro

325 330 335

Leu Phe Gly Asn Ala Gly Asn Ala Ala Pro Val Leu Val Ser Leu

340 345 350

Thr Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg

355 360 365

Ile Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp

370 375 380

Gly Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr

385 390 395 400

Ile Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro

405 410 415

Gln Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser

420 425 430

His Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg

435 440 445

Ala Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile

450 455 460

Ile Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser

465 470 475 480

Ile Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly

485 490 495

Asp Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala

500 505 510

Tyr Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg

515 520 525

Val Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val

530 535 540

Phe Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser

545 550 555 560

Leu Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly

565 570 575

Thr Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu

580 585 590

Ser Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr

595 600 605

Ala Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val

610 615 620

Asn Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val

625 630 635 640

Thr Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser

645 650 655

Asp Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys

660 665 670

His Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn

675 680 685

Phe Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr

690 695 700

Asp Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val

705 710 715 720

Thr Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln

725 730 735

Lys Ile Asp Glu Ser Lys Leu Lys Ala Phe Thr Arg Tyr Gln Leu Arg

740 745 750

Gly Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr

755 760 765

Asn Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp

770 775 780

Pro Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg

785 790 795 800

Cys Ala Pro His Leu Glu Trp Asn Pro Asp Leu Asp Cys Ser Cys Arg

805 810 815

Asp Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile

820 825 830

Asp Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile

835 840 845

Phe Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu

850 855 860

Phe Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys

865 870 875 880

Arg Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Glu Trp Glu

885 890 895

Thr Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe

900 905 910

Val Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met

915 920 925

Ile His Ala Ala Asp Lys Arg Val His Ser Ile Arg Glu Ala Tyr Leu

930 935 940

Pro Glu Leu Ser Val Ile Pro Gly Val Asn Ala Ala Ile Phe Glu Glu

945 950 955 960

Leu Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn

965 970 975

Val Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Ser Cys Trp Asn Val

980 985 990

Lys Gly His Val Asp Val Glu Glu Gln Asn Asn Gln Arg Ser Val Leu

995 1000 1005

Val Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val

1010 1015 1020

Cys Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu

1025 1030 1035

Gly Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn

1040 1045 1050

Thr Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Ile Tyr

1055 1060 1065

Pro Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Val Asn Gln Glu

1070 1075 1080

Glu Tyr Gly Gly Ala Tyr Thr Ser Arg Asn Arg Gly Tyr Asn Glu

1085 1090 1095

Ala Pro Ser Val Pro Ala Asp Tyr Ala Ser Val Tyr Glu Glu Lys

1100 1105 1110

Ser Tyr Thr Asp Gly Arg Arg Glu Asn Pro Cys Glu Phe Asn Arg

1115 1120 1125

Gly Tyr Arg Asp Tyr Thr Pro Leu Pro Val Gly Tyr Val Thr Lys

1130 1135 1140

Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp Ile Glu Ile

1145 1150 1155

Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val Glu Leu Leu

1160 1165 1170

Leu Met Glu Glu

1175

<210> 11

<211> 3534

<212> DNA

<213> Artificial sequence

<220>

<223> Fully synthetic nucleotide sequence,

encoding BCW 003 for plant use

<220>

<221> CDS

<222> (1)..(3531)

<223> BCW 003

<400> 11

atg gac aac aac ccg aac atc aac gag tgc atc ccc tac aac tgc ctc 48

Met Asp Asn Asn Pro Asn Ile Asn Glu Cys Ile Pro Tyr Asn Cys Leu

1 5 10 15

tcc aac ccg gag gtc gag gtg ctg ggc ggc gaa agg atc gag acc ggc 96

Ser Asn Pro Glu Val Glu Val Leu Gly Gly Glu Arg Ile Glu Thr Gly

20 25 30

tac act ccc atc gac atc agc ctc agc ctg acc cag ttc ctg ctc tct 144

Tyr Thr Pro Ile Asp Ile Ser Leu Ser Leu Thr Gln Phe Leu Leu Ser

35 40 45

gag ttc gtg ccc ggc gcg ggg ttc gtt ctc ggc ctg gtc gac atc atc 192

Glu Phe Val Pro Gly Ala Gly Phe Val Leu Gly Leu Val Asp Ile Ile

50 55 60

tgg ggc atc ttc ggt ccg agc cag tgg gac gcc ttt ctc gtt cag att 240

Trp Gly Ile Phe Gly Pro Ser Gln Trp Asp Ala Phe Leu Val Gln Ile

65 70 75 80

gag cag ctg atc aac cag cgc atc gag gag ttc gcc cgc aac cag gcg 288

Glu Gln Leu Ile Asn Gln Arg Ile Glu Glu Phe Ala Arg Asn Gln Ala

85 90 95

atc tcc cgg ctg gag ggc ctc tcc aac ctg tac caa atc tac gcc gag 336

Ile Ser Arg Leu Glu Gly Leu Ser Asn Leu Tyr Gln Ile Tyr Ala Glu

100 105 110

agc ttc cgg gag tgg gaa gcc gat ccg acc aac ccc gct ctc agg gag 384

Ser Phe Arg Glu Trp Glu Ala Asp Pro Thr Asn Pro Ala Leu Arg Glu

115 120 125

gag atg cgg att cag ttc aac gac atg aac tcc gct ctc acg act gcc 432

Glu Met Arg Ile Gln Phe Asn Asp Met Asn Ser Ala Leu Thr Thr Ala

130 135 140

atc cca ctc ctc gct gtg cag aac tac caa gtg ccg ctc ctg tcc gtg 480

Ile Pro Leu Leu Ala Val Gln Asn Tyr Gln Val Pro Leu Leu Ser Val

145 150 155 160

tac gtg cag gcc gcc aat ctg cac ctc tcc gtc ctc cgg gac gtt agc 528

Tyr Val Gln Ala Ala Asn Leu His Leu Ser Val Leu Arg Asp Val Ser

165 170 175

gtg ttc ggg cag cgc tgg ggc ttc gac gcc gct acc atc aac tcc cgt 576

Val Phe Gly Gln Arg Trp Gly Phe Asp Ala Ala Thr Ile Asn Ser Arg

180 185 190

tac aac gat ctc act cgc ctc atc ggc aac tac acc gac tat gcc gtg 624

Tyr Asn Asp Leu Thr Arg Leu Ile Gly Asn Tyr Thr Asp Tyr Ala Val

195 200 205

cgc tgg tac aac act ggt ctt gag aga gtc tgg ggc ccg gac agc cgc 672

Arg Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg

210 215 220

gac tgg gtg cgc tac aac cag ttc cgg cgc gag ctg acc ctc acc gtg 720

Asp Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val

225 230 235 240

ctc gac atc gta gcc ctc ttt ccc aac tac gac tcc cgg cgc tac ccg 768

Leu Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro

245 250 255

att cgc acc gtc agc cag ctc acc agg gag atc tac acc aac cct gtg 816

Ile Arg Thr Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val

260 265 270

ctg gag aac ttc gac ggc tcc ttt cgc ggg atg gcc caa cgc ata gag 864

Leu Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu

275 280 285

cag aac atc cgc caa cct cat ctg atg gac atc ctt aat tct atc acc 912

Gln Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr

290 295 300

atc tac act gac gtt cat cgc ggg ttt aac tac tgg tcg ggc cac caa 960

Ile Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln

305 310 315 320

atc act gcg tcg ccc gtt ggt ttc tcc ggc ccg gag ttc gcg ttc cct 1008

Ile Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro

325 330 335

ctg ttc gga aac gcg ggc aat gcc gct cca ccc gta ttg gtg agc ctg 1056

Leu Phe Gly Asn Ala Gly Asn Ala Ala Pro Val Leu Val Ser Leu

340 345 350

acc ggc ctc ggc atc ttc cgt aca ctg tct agc cct ctg tac aga agg 1104

Thr Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg

355 360 365

atc att ctt ggc agc ggt ccc aat aac cag gaa ctc ttc gtg ttg gac 1152

Ile Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp

370 375 380

ggc acc gag ttc agc ttc gcc agt ctt acg acc aat ttg ccc tcc aca 1200

Gly Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr

385 390 395 400

atc tat cgc cag cgc ggt act gtg gac tcc ctt gat gtg ata cca cct 1248

Ile Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro

405 410 415

cag gac aac tct gtc cca cct cgc gcc ggt ttc tcc cac cgc ctc agc 1296

Gln Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser

420 425 430

cac gtc act atg ctg agt cag gct gcg gga gcc gtg tac acc ctt cgg 1344

His Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg

435 440 445

gct ccg acg ttt agc tgg cag cac agg agc gcg act acc acg aac atc 1392

Ala Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile

450 455 460

att gcg gct gac tcc atc act caa atc cct gcc gtt aag ggt cgc tcc 1440

Ile Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser

465 470 475 480

atc atc aac aat ggg aca gtg atc tcg gga ccg ggc ttc acc ggc ggt 1488

Ile Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly

485 490 495

gac ctg gtg agg ctg tac aac gcg gac ttc aac atc aac aac agg gcg 1536

Asp Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala

500 505 510

tac ctc gaa gtc ccg atc ttc ttc cag tcg ccc agc acg aac tat cgt 1584

Tyr Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg

515 520 525

gtc agg gtc cgg tac gcc tca acc tca tcc ctc ccg gtc gat gtg gtc 1632

Val Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val

530 535 540

ttc ggc aac atc agc cac ccg acc acg ttt ccg gct acc gcc cga tcc 1680

Phe Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser

545 550 555 560

ctg gac aat ctg caa agc aac gat ttc ggc tac att gac att gcc ggg 1728

Leu Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly

565 570 575

acg ttc ctc ccg agc ctc ggc cca tcc atc ggc atc cgg ccc atg ctc 1776

Thr Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu

580 585 590

tcc acc atc aac ctg atc gtg gat cgg ttt gag ttc atc cca gtg aca 1824

Ser Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr

595 600 605

gcc act ttc gag gct gag tcc gac cta gag cgt gct cag aag gca gtc 1872

Ala Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val

610 615 620

aat gct ctg ttt acc tcc acc aat cag ctc ggc att aag acc gat gtg 1920

Asn Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val

625 630 635 640

acc gat tac cac att gac caa gtc tca aac ctc gtt gag tgc ctc tcg 1968

Thr Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser

645 650 655

gat gag ttc tac ctt gat gag aag agg gag ctt tca gag aaa gtt aag 2016

Asp Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys

660 665 670

cac gct aag aga ctc tcg gac gaa cgc aat ctg ttg caa gat ccc aac 2064

His Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn

675 680 685

ttc aga ggg atc aac cgt cag cca gac cgg gga tgg cgc ggg tcc acg 2112

Phe Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr

690 695 700

gac atc act atc cag ggc ggt gat gac gtc ttc aag gag aac tac gtg 2160

Asp Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val

705 710 715 720

acc ctg ccg ggc acc ttt gac gaa tgc tac ccc act tac ctc tac cag 2208

Thr Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln

725 730 735

aag att gac gag tcc aag ctc aag gcg ttc aca cgc tac cag ctc agg 2256

Lys Ile Asp Glu Ser Lys Leu Lys Ala Phe Thr Arg Tyr Gln Leu Arg

740 745 750

ggt tac atc gag gac tcc caa gac ctg gaa atc tac ctg atc cgc tac 2304

Gly Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr

755 760 765

aac gct aag cac gag act gtc aac gtg ccc ggc acc ggc agc ctg tgg 2352

Asn Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp

770 775 780

ccc ttg tcc gct cag agc cca atc ggc aag tgc ggc gag ccc aac cgc 2400

Pro Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg

785 790 795 800

tgc gcg ccc cac ctg gaa tgg aac ccc gac ctc gac tgt agc tgc cgc 2448

Cys Ala Pro His Leu Glu Trp Asn Pro Asp Leu Asp Cys Ser Cys Arg

805 810 815

gac gga gag aag tgc gcg cat cac tcc cac cac ttc agc ctc gac atc 2496

Asp Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile

820 825 830

gac gtc ggt tgc acc gac ctt aac gag gat ctg ggc gtt tgg gtg atc 2544

Asp Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile

835 840 845

ttc aag atc aag act cag gac ggc cac gcc cgc ctg gga aac ctg gag 2592

Phe Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu

850 855 860

ttc ctg gag gag aag ccc ctc gtt ggc gag gcc ctg gcc cgc gtc aag 2640

Phe Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys

865 870 875 880

agg gcc gag aag aaa tgg cgc gac aag cgc gag aag ctg gag tgg gag 2688

Arg Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Glu Trp Glu

885 890 895

acc aac atc gtg tac aag gaa gcg aag gag tca gtt gac gcc ctg ttc 2736

Thr Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe

900 905 910

gtc aac agc cag tac gac cag ctc cag gca gac aca aac atc gct atg 2784

Val Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met

915 920 925

atc cat gcg gcc gac aag cgc gtc cac tcc atc cgc gag gcg tac ctg 2832

Ile His Ala Ala Asp Lys Arg Val His Ser Ile Arg Glu Ala Tyr Leu

930 935 940

ccc gag ctg tcc gtc atc ccc ggc gtc aac gcc gcg atc ttt gag gag 2880

Pro Glu Leu Ser Val Ile Pro Gly Val Asn Ala Ala Ile Phe Glu Glu

945 950 955 960

ctg gag ggc cgc atc ttc acc gcc ttc tcc ctc tac gac gca cgc aac 2928

Leu Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn

965 970 975

gtt atc aag aat ggc gac ttc aac aac ggg ctg tcc tgc tgg aat gtc 2976

Val Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Ser Cys Trp Asn Val

980 985 990

aag ggc cac gtg gac gtc gag gag cag aac aac cag cgc tca gtc ctg 3024

Lys Gly His Val Asp Val Glu Glu Gln Asn Asn Gln Arg Ser Val Leu

995 1000 1005

gtc gtc ccg gag tgg gag gcc gaa gtc agc cag gaa gtc cgc gtc 3069

Val Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val

1010 1015 1020

tgc cct gga cgc ggg tac atc ctg cgc gtc act gcc tac aag gaa 3114

Cys Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu

1025 1030 1035

ggc tac gga gag ggc tgc gtc acc atc cat gag atc gaa aac aac 3159

Gly Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn

1040 1045 1050

acg gat gag ctt aag ttc agc aac tgt gtt gaa gag gaa atc tac 3204

Thr Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Ile Tyr

1055 1060 1065

ccg aac aac acg gtc acc tgc aat gat tac acc gtc aac cag gag 3249

Pro Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Val Asn Gln Glu

1070 1075 1080

gaa tac ggt gga gct tac acc tcc cgc aac agg ggc tac aac gag 3294

Glu Tyr Gly Gly Ala Tyr Thr Ser Arg Asn Arg Gly Tyr Asn Glu

1085 1090 1095

gca ccc tct gtc ccg gcc gac tac gct tca gtc tac gaa gag aag 3339

Ala Pro Ser Val Pro Ala Asp Tyr Ala Ser Val Tyr Glu Glu Lys

1100 1105 1110

tcg tac acc gac gga cgc aga gag aac ccg tgt gag ttc aac cgc 3384

Ser Tyr Thr Asp Gly Arg Arg Glu Asn Pro Cys Glu Phe Asn Arg

1115 1120 1125

ggc tac cgc gat tac acc ccg ctg cct gtc ggg tac gtc acc aaa 3429

Gly Tyr Arg Asp Tyr Thr Pro Leu Pro Val Gly Tyr Val Thr Lys

1130 1135 1140

gag ctg gaa tac ttc cca gag acc gac aaa gtc tgg att gag atc 3474

Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp Ile Glu Ile

1145 1150 1155

ggc gag acc gag ggc acg ttc atc gtg gac tcc gtc gaa ctc ctt 3519

Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val Glu Leu Leu

1160 1165 1170

ctg atg gaa gag tga 3534

Leu Met Glu Glu

1175

<210> 12

<211> 1177

<212> PROTEIN

<213> Artificial sequence

<220>

<223> Synthetic construction

<400> 12

Met Asp Asn Asn Pro Asn Ile Asn Glu Cys Ile Pro Tyr Asn Cys Leu

1 5 10 15

Ser Asn Pro Glu Val Glu Val Leu Gly Gly Glu Arg Ile Glu Thr Gly

20 25 30

Tyr Thr Pro Ile Asp Ile Ser Leu Ser Leu Thr Gln Phe Leu Leu Ser

35 40 45

Glu Phe Val Pro Gly Ala Gly Phe Val Leu Gly Leu Val Asp Ile Ile

50 55 60

Trp Gly Ile Phe Gly Pro Ser Gln Trp Asp Ala Phe Leu Val Gln Ile

65 70 75 80

Glu Gln Leu Ile Asn Gln Arg Ile Glu Glu Phe Ala Arg Asn Gln Ala

85 90 95

Ile Ser Arg Leu Glu Gly Leu Ser Asn Leu Tyr Gln Ile Tyr Ala Glu

100 105 110

Ser Phe Arg Glu Trp Glu Ala Asp Pro Thr Asn Pro Ala Leu Arg Glu

115 120 125

Glu Met Arg Ile Gln Phe Asn Asp Met Asn Ser Ala Leu Thr Thr Ala

130 135 140

Ile Pro Leu Leu Ala Val Gln Asn Tyr Gln Val Pro Leu Leu Ser Val

145 150 155 160

Tyr Val Gln Ala Ala Asn Leu His Leu Ser Val Leu Arg Asp Val Ser

165 170 175

Val Phe Gly Gln Arg Trp Gly Phe Asp Ala Ala Thr Ile Asn Ser Arg

180 185 190

Tyr Asn Asp Leu Thr Arg Leu Ile Gly Asn Tyr Thr Asp Tyr Ala Val

195 200 205

Arg Trp Tyr Asn Thr Gly Leu Glu Arg Val Trp Gly Pro Asp Ser Arg

210 215 220

Asp Trp Val Arg Tyr Asn Gln Phe Arg Arg Glu Leu Thr Leu Thr Val

225 230 235 240

Leu Asp Ile Val Ala Leu Phe Pro Asn Tyr Asp Ser Arg Arg Tyr Pro

245 250 255

Ile Arg Thr Val Ser Gln Leu Thr Arg Glu Ile Tyr Thr Asn Pro Val

260 265 270

Leu Glu Asn Phe Asp Gly Ser Phe Arg Gly Met Ala Gln Arg Ile Glu

275 280 285

Gln Asn Ile Arg Gln Pro His Leu Met Asp Ile Leu Asn Ser Ile Thr

290 295 300

Ile Tyr Thr Asp Val His Arg Gly Phe Asn Tyr Trp Ser Gly His Gln

305 310 315 320

Ile Thr Ala Ser Pro Val Gly Phe Ser Gly Pro Glu Phe Ala Phe Pro

325 330 335

Leu Phe Gly Asn Ala Gly Asn Ala Ala Pro Val Leu Val Ser Leu

340 345 350

Thr Gly Leu Gly Ile Phe Arg Thr Leu Ser Ser Pro Leu Tyr Arg Arg

355 360 365

Ile Ile Leu Gly Ser Gly Pro Asn Asn Gln Glu Leu Phe Val Leu Asp

370 375 380

Gly Thr Glu Phe Ser Phe Ala Ser Leu Thr Thr Asn Leu Pro Ser Thr

385 390 395 400

Ile Tyr Arg Gln Arg Gly Thr Val Asp Ser Leu Asp Val Ile Pro Pro

405 410 415

Gln Asp Asn Ser Val Pro Pro Arg Ala Gly Phe Ser His Arg Leu Ser

420 425 430

His Val Thr Met Leu Ser Gln Ala Ala Gly Ala Val Tyr Thr Leu Arg

435 440 445

Ala Pro Thr Phe Ser Trp Gln His Arg Ser Ala Thr Thr Thr Asn Ile

450 455 460

Ile Ala Ala Asp Ser Ile Thr Gln Ile Pro Ala Val Lys Gly Arg Ser

465 470 475 480

Ile Ile Asn Asn Gly Thr Val Ile Ser Gly Pro Gly Phe Thr Gly Gly

485 490 495

Asp Leu Val Arg Leu Tyr Asn Ala Asp Phe Asn Ile Asn Asn Arg Ala

500 505 510

Tyr Leu Glu Val Pro Ile Phe Phe Gln Ser Pro Ser Thr Asn Tyr Arg

515 520 525

Val Arg Val Arg Tyr Ala Ser Thr Ser Ser Leu Pro Val Asp Val Val

530 535 540

Phe Gly Asn Ile Ser His Pro Thr Thr Phe Pro Ala Thr Ala Arg Ser

545 550 555 560

Leu Asp Asn Leu Gln Ser Asn Asp Phe Gly Tyr Ile Asp Ile Ala Gly

565 570 575

Thr Phe Leu Pro Ser Leu Gly Pro Ser Ile Gly Ile Arg Pro Met Leu

580 585 590

Ser Thr Ile Asn Leu Ile Val Asp Arg Phe Glu Phe Ile Pro Val Thr

595 600 605

Ala Thr Phe Glu Ala Glu Ser Asp Leu Glu Arg Ala Gln Lys Ala Val

610 615 620

Asn Ala Leu Phe Thr Ser Thr Asn Gln Leu Gly Ile Lys Thr Asp Val

625 630 635 640

Thr Asp Tyr His Ile Asp Gln Val Ser Asn Leu Val Glu Cys Leu Ser

645 650 655

Asp Glu Phe Tyr Leu Asp Glu Lys Arg Glu Leu Ser Glu Lys Val Lys

660 665 670

His Ala Lys Arg Leu Ser Asp Glu Arg Asn Leu Leu Gln Asp Pro Asn

675 680 685

Phe Arg Gly Ile Asn Arg Gln Pro Asp Arg Gly Trp Arg Gly Ser Thr

690 695 700

Asp Ile Thr Ile Gln Gly Gly Asp Asp Val Phe Lys Glu Asn Tyr Val

705 710 715 720

Thr Leu Pro Gly Thr Phe Asp Glu Cys Tyr Pro Thr Tyr Leu Tyr Gln

725 730 735

Lys Ile Asp Glu Ser Lys Leu Lys Ala Phe Thr Arg Tyr Gln Leu Arg

740 745 750

Gly Tyr Ile Glu Asp Ser Gln Asp Leu Glu Ile Tyr Leu Ile Arg Tyr

755 760 765

Asn Ala Lys His Glu Thr Val Asn Val Pro Gly Thr Gly Ser Leu Trp

770 775 780

Pro Leu Ser Ala Gln Ser Pro Ile Gly Lys Cys Gly Glu Pro Asn Arg

785 790 795 800

Cys Ala Pro His Leu Glu Trp Asn Pro Asp Leu Asp Cys Ser Cys Arg

805 810 815

Asp Gly Glu Lys Cys Ala His His Ser His His Phe Ser Leu Asp Ile

820 825 830

Asp Val Gly Cys Thr Asp Leu Asn Glu Asp Leu Gly Val Trp Val Ile

835 840 845

Phe Lys Ile Lys Thr Gln Asp Gly His Ala Arg Leu Gly Asn Leu Glu

850 855 860

Phe Leu Glu Glu Lys Pro Leu Val Gly Glu Ala Leu Ala Arg Val Lys

865 870 875 880

Arg Ala Glu Lys Lys Trp Arg Asp Lys Arg Glu Lys Leu Glu Trp Glu

885 890 895

Thr Asn Ile Val Tyr Lys Glu Ala Lys Glu Ser Val Asp Ala Leu Phe

900 905 910

Val Asn Ser Gln Tyr Asp Gln Leu Gln Ala Asp Thr Asn Ile Ala Met

915 920 925

Ile His Ala Ala Asp Lys Arg Val His Ser Ile Arg Glu Ala Tyr Leu

930 935 940

Pro Glu Leu Ser Val Ile Pro Gly Val Asn Ala Ala Ile Phe Glu Glu

945 950 955 960

Leu Glu Gly Arg Ile Phe Thr Ala Phe Ser Leu Tyr Asp Ala Arg Asn

965 970 975

Val Ile Lys Asn Gly Asp Phe Asn Asn Gly Leu Ser Cys Trp Asn Val

980 985 990

Lys Gly His Val Asp Val Glu Glu Gln Asn Asn Gln Arg Ser Val Leu

995 1000 1005

Val Val Pro Glu Trp Glu Ala Glu Val Ser Gln Glu Val Arg Val

1010 1015 1020

Cys Pro Gly Arg Gly Tyr Ile Leu Arg Val Thr Ala Tyr Lys Glu

1025 1030 1035

Gly Tyr Gly Glu Gly Cys Val Thr Ile His Glu Ile Glu Asn Asn

1040 1045 1050

Thr Asp Glu Leu Lys Phe Ser Asn Cys Val Glu Glu Glu Ile Tyr

1055 1060 1065

Pro Asn Asn Thr Val Thr Cys Asn Asp Tyr Thr Val Asn Gln Glu

1070 1075 1080

Glu Tyr Gly Gly Ala Tyr Thr Ser Arg Asn Arg Gly Tyr Asn Glu

1085 1090 1095

Ala Pro Ser Val Pro Ala Asp Tyr Ala Ser Val Tyr Glu Glu Lys

1100 1105 1110

Ser Tyr Thr Asp Gly Arg Arg Glu Asn Pro Cys Glu Phe Asn Arg

1115 1120 1125

Gly Tyr Arg Asp Tyr Thr Pro Leu Pro Val Gly Tyr Val Thr Lys

1130 1135 1140

Glu Leu Glu Tyr Phe Pro Glu Thr Asp Lys Val Trp Ile Glu Ile

1145 1150 1155

Gly Glu Thr Glu Gly Thr Phe Ile Val Asp Ser Val Glu Leu Leu

1160 1165 1170

Leu Met Glu Glu

1175

<---

Claims (36)

1. A polynucleotide construct for imparting resistance to an insect of the Lepidoptera order, containing a nucleotide sequence encoding: a) an insecticidal protein having an amino acid sequence comprising amino acids 1 to 607 of a sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO : 10 and SEQ ID NO: 12; or b) an insecticidal protein having the amino acid sequence from position 7 to 607 as specified in any of SEQ ID NOs: 2, 4, 6, 8, 10 or 12, wherein said nucleotide sequence is operably linked to a heterologous promoter sequence. 2. A protein toxic to the black cutworm of the order Lepidoptera, containing: a) amino acid sequence as indicated in SEQ ID NO: 2 from positions 256 to 606; b) amino acid sequence as indicated in SEQ ID NO: 4 from position 257 to 607; and c) amino acid sequence as indicated in SEQ ID NO: 6 from positions 257 to 607. 3. A polynucleotide construct according to claim 1, characterized in that said insecticidal protein is active against an insect of the order Lepidoptera, selected from the group consisting of: Spodoptera frugiperda , Spodoptera exigua , Spodoptera litura , Mamestra configurata , Striacosta albicosta , Trichoplusia ni , Pseudoplusia includens , Anticarsia gemmatalis , Hypena scabra , Heliothis virescens , Agrotis subterranea , Pseudaletia unipuncta , Agrotis ipsilon , Agrotis orthogonia, Ostrinia nubilalis , Amyelois transitella , Crambus caliginosellus , Herpetogramma licarsisalis , Homoeosoma electellum , Elasmopalpus lignosellus , Cydia pomonella , Endopiza viteana , Grapholita molesta , Suleima helianthana , Plutella xylostella , Pectinophora gossypiella , Lymantria dispar , Alabama argillacea , Archips argyrospila , Archips rosana , Chilo suppressalis , Cnaphalocrocis medinalis , Crambus caliginosellus , Crambus teterrellus , Diatraea grandiosella , Diatraea insularis saccharalis , a , Egrias vittella , Helicoverpa armigera , Helicoverpa zea , Heliothis virescens , Herpetogramma licarsisalis , Lobesia botrana , Pectinophora gossypiella , Phyllocnistis citrella , Pieris brassicae , Pieris rapae , Plutella xylostella , and Tuta absoluta . 4. Protein according to claim 2, additionally containing bioactivity against Lepidoptera species selected from the group consisting of: Spodoptera frugiperda , Spodoptera exigua , Spodoptera litura , Mamestra configurata , Striacosta albicosta , Trichoplusia ni , Pseudoplusia includens , Anticarsia gemmatalis , Hypena scabra , Heliothis virescens , Agrotis subterranea , Pseudaletia unipuncta , Agrotis ipsilon , Agrotis orthogonia, Ostrinia nubilalis , Amyelois transitella , Crambus caliginosellus , Herpetogramma licarsisalis , Homoeosoma electellum , Elasmopalpus lignosellus , Cydia pomonella , Endopiza viteana , Grapholita molesta , Suleima helianthana , Plutella xylostella , Pectinophora gossypiella , Lymantria dispar , Alabama argillacea , Archips argyrospila , Archips rosana , Chilo suppressalis , Cnaphalocrocis medinalis , Crambus caliginosellus , Crambus teterrellus , Diatraea grandiosella , Diatraea saccharalis , Earias insulana , Egrias vittella , Helicoverpa armigera liothis virescens , Herpetogramma licarsisalis , Lobesia botrana , Pectinophora gossypiella , Phyllocnistis citrella , Pieris brassicae , Pieris rapae , Plutella xylostella , and Tuta absoluta . 5. A vector expressing a polynucleotide construct according to claim 1. 6. A host cell having resistance to an insect of the Lepidoptera order, containing a polynucleotide construct according to claim 1, characterized in that said host cell is selected from the group consisting of a bacterial cell, a yeast cell and a plant cell. 7. Host cell according to claim 6, characterized in that said plant cell is selected from the group consisting of a plant cell: alfalfa, banana, barley, beans, broccoli, cabbage, ornamental cabbage, carrots, cassava, castor bean, cauliflower, celery, chickpeas, bok choy, citrus, coconut, coffee, corn, clover, cotton, cucurbits, cucumber, menzies, eggplant, eucalyptus, flax, garlic, grapes, hops, leek, lettuce, frankincense pine, millet, melon, walnut, oats, olive tree, onion, ornamental plants, palm, pasture grasses, pea, peanut, pepper, pigeon pea, pine, potato, poplar, pumpkin, radiant pine, radish, rapeseed, rice, rhizomes, rye, wild saffron, shrub, sorghum, southern pine, soybean, spinach, strawberry, sugar beet, sugar cane, sunflower, sugar corn, amber, sweet potato, switchgrass, tea, tobacco, tomato, triticale, turfgrass, watermelon and wheat. 8. A transgenic plant having resistance to an insect of the order Lepidoptera, where said plant contains a polynucleotide construct according to claim 1. 9. A seed having resistance to an insect of the order Lepidoptera and wherein said seed contains a detectable amount of said polynucleotide construct. 10. A transgenic plant according to claim 8, characterized in that seeds, pollen, progeny, plant cells, plant tissue and commercial products obtained from said plant contain a detectable amount of said polynucleotide construct. 11. A biological sample having resistance to an insect of the Lepidoptera order, where the biological sample contains a detectable amount of the polynucleotide construct according to claim 1. 12. Composition for controlling insects of the Lepidoptera order, containing an insecticidally effective amount of a protein according to claim 2, and: a) an agent other than said protein and also toxic to the insect order Lepidoptera, said agent being selected from the group consisting of a polypeptide having an amino acid sequence different from said protein, an RNA molecule and a chemical compound; or b) an agent selected from the group consisting of: Cry1A, Cry1Ab, Cry1Ac, Cry1A.105, Cry1Ae, Cry1B, Cry1C, Cry1D, Cry1E, Cry1F, Cry1G, Cry1H, Cry1I, Cry1J, Cry1K, Cry1L, Cry2A, Cry2Ab, Cry2Ae, Cry4B, Cry6, Cry7, Cry8, Cry9, Cry15, Cry43A, Cry43B, ET35, ET66, TIC400, TIC800, TIC807, TIC834, TIC853, TIC1415, VIP3A, VIP3Ab, AXMI insecticidal proteins, DIG insecticidal proteins, eHIPs and VIP proteins. 13. Composition according to claim 12, additionally containing an additional pesticidal agent, and the specified additional agent is selected from the group consisting of: Cry1C, Cry3A, Cry3B, Cry34, Cry35, Cry51Aa1, ET29, ET33, ET34, ET70, TIC407, TIC417, TIC431 , TIC901, TIC1201, TIC3131, 5307, DIG-10, Axmi184, Axmi205 and AxmiR1. 14. A method for obtaining seeds containing a polynucleotide sequence according to claim 1, and this method includes: a) transforming one or more seeds with a vector according to claim 5, b) planting one or more seeds containing said polynucleotide; c) growing plants from said seed; and d) harvesting seeds from said plants, said seed containing said polynucleotide construct. 15. The plant according to claim 8, additionally containing: a) a transgenic corn plant selected from the group consisting of: DKB89614-9, MON801, MON802, MON809, MON810, MON863, MON88017, MON89034, transformant 4114-3, transformant 5307, DAS59122-7, Bt10, Bt11, Bt176, CBH -351, DKB-83614-9, MIR162, MIR604, TC1507, TC6275, transform 676, transform 678, transform 680, transform 98140, DAS40278-9, DKB89790-5, MON21-9, HCEM485, MON832, MON87427, NK603, T14 , T25 and VCO01981-5; b) a transgenic soybean plant selected from the group consisting of: MON87751, DAS81419-2, MON87701, A2704-12, A2704-21, A5547-127, A5547-35, CV127, DAS44406-6, DAS68416-4, DP356043, FG72 , MON4032, ACS-GM003-1, MON87705, MON87708, MON89788, W62, W98 and GFM Cry1A; c) a transgenic cotton plant selected from the group consisting of: DAS24236-5, DAS21023-5, transformant 31707, transformant 31803, transformant 31807, transformant 31808, transformant 42317, BNLA-601, COT102, COT67B, transformant 1, GHB119, GK12 , MON15985, MLS9124, MON1076, MON531, MON757, T303-3, T304-40, SGK321, transformant 19-51a, GHB614, LLCotton25, MON88701, MON88702, MON1445, MON1698 and MON88913; d) transgenic sugar cane plant transformant NXI-1T; and e) a transgenic rice plant selected from the group consisting of: LLRICE06, LLRICE601, LLRIC2E62, GM-A17054 and GM-A17054. 16. Toxic for Lepidoptera protein containing in a functional relationship: a) a first peptide segment containing the amino acid sequence of Cry1A domain I; b) a second peptide segment containing residues 255-606 of SEQ ID NO: 2; wherein said Lepidoptera toxic protein exhibits toxic bioactivity when tested against Lepidoptera pests selected from the group consisting of: epsilon cutworm ( A. ipsilon ), cotton cutworm ( H. zea) , Striacosta albicosta ( S. albicosta) , corn moth ( O. nubilalis ), southwestern corn moth ( D. grandiosella) , cabbage cutworm ( T. ni) , soybean cutworm ( P. includens) , grass cutworm ( S. frugiperda) and Diatraea saccharalis ( D. saccharalis) .

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