CN1174093C - Polypeptide having cold-resistant pyruvate phosphate dikinase activity, DNA coding for the polypeptide, recombinant vector containing the DNA and transformed plant - Google Patents

Abstract

A novel polypeptide having a cold-resistant pyruvate phosphate dikinase activity as a means for imparting cold resistance to plants; a cloned DNA coding for the polypeptide; and a recombinant vector containing the DNA. The polypeptide has an amino acid sequence, in any of the following dikinase (1) and polypeptide (2), wherein at least one amino acid residue present in the region with a length of one-sixth of the total length from the C-terminus of the amino acid sequence has been replaced by another amino acid residue and also has a cold-resistant pyruvate phosphate dikinase activity: (1) a pyruvate phosphate dikinase having an amino acid sequence represented by any of the SEQ ID Nos. 1 to 4 in the Sequence Listing, and (2) a polypeptide having an amino acid sequence which is at least 50 % homologous with the amino acid sequence of the dikinase (1) and also having a pyruvate phosphate dikinase activity.

Description

Polypeptide and corresponding DNA, recombinant vectors and transformed plant cell that the two kinase activities of low temperature resistant pyruvate phosphate are arranged

Technical field

The present invention relates to as give the means of plant with lower temperature resistance, have an active polypeptide of novel cold-resistant pyruvate phosphate dikinase (hereinafter to be referred as " PPDK "), with the clone DNA of its coding with contain the recombinant vectors of this DNA.In addition, also relate to the plant of transition with the invention described above DNA.

Background technology

C ₄Plant is at high light, high temperature, low CO ₂Condition under high photosynthesis ability is arranged, but can adapt to the cold condition except that a part, generally reduce greatly in low temperature time resultant quantity.PPDK (EC 2.7.9.1, energy catalysis generates the reaction of AMP, phosphoenolpyruvic acid and tetra-sodium from ATP, pyruvic acid and ortho-phosphoric acid) is C ₄One of important enzyme of approach, but its activity to compare with the photosynthesis speed of leaf texture be not enough, thereby become C ₄One of enzyme of restriction carbon dioxide fixation speed in the photosynthesis.In addition, when it is found that PPDK, just point out that it is the low-temperature sensitive enzyme.Under the situation of corn PPDK, the interior curved point of enzymic activity is 11.7 ℃, and this temperature is consistent with the growth ultimate temperature of corn.From these facts, can think that PPDK is C ₄One of reason that plant low temperature light compositing speed is low by improving the low-temperature sensitive of PPDK, perhaps can reduce this C of corn ₄The growth ultimate temperature of plant.The pale brown colored chrysanthemum of feverfew ( Flaveria Brownii) belong to C ₃/ C ₄The osculant classification, even known its PPDK also hardly can inactivation (Burnell JN:A comparative study of the cold-sensitivityof pyruvate when 0 ℃ subzero treatment, Pi dikinase in Flaveria species.Plant CellPhysiol, 31,295-297 (1990)).

If make gene cloneization with the low temperature resistant PPDK coding of above-mentioned pale brown colored chrysanthemum, with it plant is made the transition again, then be expected to give plant with lower temperature resistance.

Disclosure of the Invention

Therefore, the purpose of this invention is to provide as give the means of plant with lower temperature resistance, have novel can the active polypeptide of low temperature resistant PPDK, with the clone DNA of its coding and the recombinant vectors that contains this DNA.In addition, purpose of the present invention also provides the plant that makes the transition with above-mentioned DNA of the present invention.

The result that people such as the application's inventor further investigate, successfully make the full PPDK gene cloneization of pale brown colored chrysanthemum and determined its base sequence and its amino acid sequence coded, in addition, also successfully in this PPDK gene, identify the zone of giving lower temperature resistance, thereby finished the present invention.

Promptly, the invention provides in following (1), (2) in any one, have counting that at least more than one radical amino acid replacement becomes the aminoacid sequence of other amino-acid residue in the scope that accounts for total length 1/6, and have the polypeptide of cold-resistant pyruvate phosphate dikinase activity from the C of aminoacid sequence end:

(1) has the two kinases of pyruvate phosphate of sequence table sequence number 1 to 4 aminoacid sequence shown in any one;

(2) have and the identical aminoacid sequence and have the polypeptide of the two kinase activities of pyruvate phosphate more than 50% of aminoacid sequence shown in (1).

In addition, the present invention also provides and will have the clone DNA that the present invention can the active peptide coding of low temperature resistant PPDK.And the present invention also provides and contains DNA of the present invention, can express the recombinant vectors with polypeptide that can low temperature resistant PPDK in the host.And then the present invention also provides the plant that makes the transition with DNA of the present invention.

The present invention makes the PPDK gene cloneization with lower temperature resistance, and has determined its base sequence.And, also determined to give in this gene the zone of lower temperature resistance.Therefore, by make plant transition with gene of the present invention, can make the PPDK of low-temperature sensitive become lower temperature resistance with low-temperature sensitive PPDK.In addition, be combined to the same section of low-temperature sensitive PPDK, also can make low-temperature sensitive PPDK become lower temperature resistance by making above-mentioned zone of giving lower temperature resistance.By doing like this, also might cultivate this plant at the plug cryogenic region that can not cultivate this plant originally even make.Therefore, the present invention is expected agricultural is made huge contribution.

The accompanying drawing simple declaration

Fig. 1 is a width of cloth mode chart, and the construction process of the expression vector that contains one of PPDK gene of the present invention example is described.

Fig. 2 is PPDK when 0 ℃ the preserving enzymic activity time dependent figure of explanation with pale brown colored chrysanthemum, bidentate chrysanthemum chrysanthemum (Flaveriabidentis) and the corn of colibacillus expression.

Fig. 3 is a width of cloth mode chart, and the construction process of the chimeric gene of corn PPDK and pale brown colored chrysanthemum PPDK is described.

Implement optimal morphology of the present invention

According to the present invention, make pale brown colored chrysanthemum PPDK gene cloneization, and determined its base sequence and the reckoning aminoacid sequence of its coding with lower temperature resistance.This base sequence and aminoacid sequence are represented with the sequence number 5 of sequence table.This sequence is as being described in detail in following examples, be that greenery from pale brown colored chrysanthemum extract full RNA, press well-established law and make the cDNA storehouse, plaque hydridization is carried out as the probe in high homology zone with the PPDK gene base sequence (sequence number 2) of bidentate chrysanthemum chrysanthemum (Flaveria bidentis) (sequence number 1) and corn, select positive clone carry out cloneization, determine base sequence with dideoxy method again and definite.This sequence has and belongs to the same high homology of pale brown colored chrysanthemum PPDK gene together, also have the homology more taller than corn PPDK, but also with in full accord from N-terminal sequence, C-terminal sequence and the internal sequence of the direct purified PPDK of pale brown colored chrysanthemum greenery, thereby, obviously be the PPDK gene of pale brown colored chrysanthemum.And the PPDK gene order of bidentate chrysanthemum chrysanthemum is also carried out plaque hydridization, is selected positive clone to carry out cloneization to determine with dideoxy method again as the probe with corn cDNA.

Sequence number with shown in aminoacid sequence be novel, compare with the PPDK aminoacid sequence that belongs to bidentate chrysanthemum chrysanthemum together, 40 amino-acid residue differences are arranged.And, compare with the PPDK aminoacid sequence of corn, about 180 amino-acid residue differences are also arranged.Pale brown colored chrysanthemum PPDK aminoacid sequence shown in the sequence number 5, especially with belong to bidentate chrysanthemum chrysanthemum PPDK aminoacid sequence together and compare, although so high homology is arranged, but the PPDK of bidentate chrysanthemum chrysanthemum is a low-temperature sensitive, the PPDK of pale brown colored chrysanthemum then is low temperature resistant, and this shows that the fine difference of aminoacid sequence has caused the significant differences on the proterties.The invention provides amino acid sequences encoded clone PPDK gene shown in this sequence number 5.As mentioned above, this aminoacid sequence is novel, and has and have the such unusual effect of lower temperature resistance.Gene of the present invention is not limited to the gene of base sequence shown in the sequence number 5, so long as this amino acid sequence encode also can be able to be had any base sequence.

All inventors of the application also once attempted shown in the identification sequence number 5 zone relevant with giving lower temperature resistance in the pale brown colored chrysanthemum PPDK gene.Promptly, as describing in detail in following examples, with restriction enzyme the PPDK gene of pale brown colored chrysanthemum is cut into three impartial parts of size, the exchange of the corresponding zone of zone of cutting apart and corn PPDK gene is formed mosaic PPDK gene, determine whether the PPDK of this chimeric gene coding has lower temperature resistance.Results verification exists the field of giving lower temperature resistance in last 1/3 zone of pale brown colored chrysanthemum.In addition, this last 1/3 zone further cuts into two five equilibriums with restriction enzyme again, uses with quadrat method to confirm whether there is the zone of giving lower temperature resistance in its any part.Its results verification in the downstream, XhoI position of pale brown colored chrysanthemum PPDK gene shown in the sequence number 5, promptly in aminoacid sequence shown in the sequence number 5 from the aminoacid sequence of No. 955 Xie Ansuans of No. 832 arginine to the (being " giving the lower temperature resistance sequence ") to call this sequence in the following text, exist the function of giving lower temperature resistance.

That is, having distinguished that the PPDK zone relevant with lower temperature resistance is present in from C-terminal accounts in the scope of total length 1/6.On the other hand, in sequence table sequence number 1, list the base sequence of bidentate chrysanthemum chrysanthemum PPDK encoding gene and the aminoacid sequence of deduction, and in sequence number 2, listed the base sequence of corn PPDK encoding gene and aminoacid sequence (the Journalof Biochemistry of deduction 263, 11080-11083 (1988)).Listed in the sequence number 3 bacterium parasymbiosis bacterioide ( Bacteroides Symbiosus) the PPDK encoding gene base sequence and infer aminoacid sequence (Biochemistry 29, 10757-10765 (1990)).Listed in the sequence number 4 bacterium class histolysis inner amoeba ( Entamoeba Histolytiea) the PPDK encoding gene base sequence and infer aminoacid sequence (Molecular and Biochemical Parasitology 6 2, 153-156 (1993)).As mentioned above, give the zone of PPDK owing to distinguish count 1/6 scope that accounts for total length from C-terminal in by the present invention with lower temperature resistance, thereby become other amino-acid residue by making these aminoacid sequences shown in the sequence number 1 to 4 count at least 1 the above radical amino acid replacement that accounts in total length 1/6 scope from C-terminal, might obtain lower temperature resistance PPDK.In addition, the what is called here " lower temperature resistance " mean its activity when enzyme is placed 20 minutes under 0 ℃ temperature be before placing more than 60%.

As mentioned above, shown in the sequence number 5 in the aminoacid sequence zone from No. 955 Xie Ansuans of No. 832 arginine to the determined lower temperature resistance, thereby the corresponding part of low-temperature sensitive PPDK described in the sequence number 1 to 4 is replaced as shown in the sequence number 5 in the aminoacid sequence from the aminoacid sequence of No. 955 Xie Ansuans of No. 832 arginine to the, can make its low-temperature sensitive PPDK become lower temperature resistance.This discovery is extremely important, because utilize this result can give desirable PPDK with lower temperature resistance.In addition, as giving PPDK method with lower temperature resistance, as described in the following Examples, be not limited to by making the corresponding part of giving lower temperature resistance sequence and low-temperature sensitive PPDK among the pale brown colored chrysanthemum PPDK exchange the method for making chimeric gene, also can make the corresponding part happening part specificity variation (site-directed mutagenesis) of the low-temperature sensitive PPDK of its value thing itself, thus change over pale brown colored chrysanthemum PPDK give the low temperature properties sequence identical sequence.Therefore, comprise and above-mentionedly give the low temperature properties sequence, have the active peptide coding clone of PPDK DNA and all belong to scope of the present invention.Specifically, No. 869 of aminoacid sequence shown in the sequence number 1 is replaced as the PPDK of proline(Pro), and the PPDK that is replaced as leucine and Xie Ansuan for No. 885 and No. 952 respectively, and lower temperature resistance is all arranged.

The PPDK that gives lower temperature resistance is not limited to those shown in the sequence table 1 to 4, so long as have its separately homology 50% above person get final product.Being preferably can be more than 48.5% with the homology of pale brown colored chrysanthemum PPDK base sequence, and being more preferably can be more than 90%.

In general, even under the situation of the aminoacid sequence generation minor alteration of the peptide that physiologically active is arranged, promptly, even there are one or more amino acid to be replaced or lack or increased under one or more amino acid whose situations in this aminoacid sequence, the still retainable situation of the physiologically active of this peptide is a well known fact.Therefore, have and carry out still having the active polypeptide of lower temperature resistance PPDK after such modification in the polypeptide of aminoacid sequence shown in the sequence number 5 and also belong to scope of the present invention.That is, in aminoacid sequence shown in the sequence number 5, addition, disappearance or replaced one or more amino acid after still have the active polypeptide of lower temperature resistance PPDK and also belong to scope of the present invention.Equally, in base sequence shown in the sequence number 5, addition, disappearance or replaced one or more Nucleotide after still have the active peptide coding DNA of lower temperature resistance PPDK and also belong to this

Scope of invention.

Cause that amino acid addition, disappearance or metathetical DNA can be such as (for example making a variation this well-known technology by means of bringing out site specific, Nucleic Acid Research, Vol.10, No.20, p 6487-6500,1982) implement, in this specification sheets so-called " one or more amino acid " mean can be by means of site specific make a variation provocation method addition, disappearance or metathetical amino acid number.

Bringing out of site specific variation can be such as utilizing single stranded phage DNA complementary synthetic oligonucleotide primer specific inconsistent and that should be made a variation except that desired variation to be performed as follows.That is, in phage, synthesize complementary strand as introduction, but make the host bacteria transition of carrier band phage again with resulting double-stranded DNA with above-mentioned synthetic oligonucleotide.The culture of bacterium is coated on the agar transition, from containing the unicellular formation plaque of phage.If carry out like this, in theory, 50% new bacterium colony contains as strand, the phage of variation is arranged, and all the other 50% contains original sequence.Then, make resulting plaque at an energy with the on all four DNA hydridization of the DNA with above-mentioned desired variation but the discord and the temperature of the inconsistent DNA hydridization of original chain with the synthetic probe that kinases is handled hydridization take place.Collect, cultivate the plaque with this probe's hydridization then, reclaim DNA.

In addition, as in the aminoacid sequence of enzyme, replacing, lack or insert the method that one or more amino acid do not make it to lose enzymic activity again, except that above-mentioned site specific variation provocation method, also useful mutagen is handled the method for gene, and make that gene generation selectivity ruptures, removes then, addition or replace the method that selected Nucleotide connects again.

By with the PPDK gene of pale brown colored chrysanthemum or contain it and give the lower temperature resistance sequence and have the active DNA of PPDK and make plant transition, can obtain the plant of lower temperature resistance.At this moment, as the better example of transformed plant, can enumerate corn, sugarcane, millet (grain), barnyard grass and jowar etc., but be not limited to these.

Method transition of plant is sophisticated, better adopt based on the swelling edaphic bacillus ( Agrobacterium Tumefaciens) method.The plant transition method of utilizing the swelling edaphic bacillus is well-known in the art, and in this way, no matter dicotyledons (for example the spy opens flat 4-330234 communique) still is a monocotyledons (WO 94/00977) can both make the transition.Perhaps, can also import in the protoplasma of plants, and make DNA go up, squeeze into the medium way of embryo of plant, also can make the transition attached to tungsten particle etc. with common method such as electroporation method etc.The concrete grammar of these transition is narrated in following examples.

Embodiment

Following with reference to implementing to be described more specifically the present invention.Certainly, the invention is not restricted to following embodiment.

1. determining of the cloneization of pale brown colored chrysanthemum PPDK gene and base sequence

(1) cloneization of the making in cDNA storehouse and full-length cDNA

(i) making in cDNA storehouse

Greenery (60g) from pale brown colored chrysanthemum (F.brownii) separate full RNA with Guanidinium hydrochloride/phynol method.Can obtain 26.5mg RNA at the lithium post precipitation in this way.With the pillar of having filled oligo dT Cellulose Type 7 (Pharmacia companies), obtain 118.9 μ g poly A (+) RNA from 13.2mg RNA then by common method.In the preparation in cDNA storehouse, TimeSaver cDNA Synthesis kit (Pharmacia company), Lambda ZAPII carrier (Stratagene company) and the incidental filler reagent of the λ gt10 of cDNA clone system (Amersham company) have been used.With the EcoRI/NotI linker dna fragmentation is inserted the EcoR I position of Lambda ZAP II carrier, thereby made the cDNA storehouse.The size in the cDNA storehouse of made is 41.5 ten thousand pfu.Also used XL1-Blue as host cell.

(ii) probe's preparation

Utilization be equivalent near the working position introduction 5 ': a GACGGCTAAAAAGAGGGT (according to the cDNA homology of the cDNA of bidentate chrysanthemum chrysanthemum PPDK and corn PPDK high partial design) and the introduction R:TATCGAGAAACCTTCTATAC (part of bidentate chrysanthemum chrysanthemum PPDK sequence, complementary strand) carrying out reverse transcription PCR enlarges pale brown colored chrysanthemum RNA, enlarge segmental pCR II carrier (can available from Invitrogen company) as template with having, carry out that PCR makes that this fragment enlarges, behind the electrophoresis, from gel, reclaim DNA with same introduction with SUPREC-01 (precious wine is made).Use this technology, can obtain the dna fragmentation of the 428bp (bp=base pair) that mature protein N-terminal downstream 24bp begins.With Multiprime dna marker system (Amersham company) this fragment is carried out ³²The P mark prepares the probe.

The (iii) cloneization of the pale brown colored chrysanthemum cDNA of total length

As the probe, screen the cDNA storehouse with above-mentioned dna fragmentation with plaque hydridization method.Use Hybond N ⁺As the hydridization strainer, the hydridization condition is to spend the night in 65 ℃ in 6 * SSC, 5 * Denhalt ' s solution, 0.1%SDS, 100 μ g/ml sex change salmon spermary DNA (Amersham company).Wash conditions is to carry out 5 minutes in room temperature with 2 * SSC, 0.1%SDS, carries out 90 minutes in room temperature with 2 * SSC, 0.1%SDS, further uses 1 * SSC, 0.1%SDS to carry out 90 minutes at 68 ℃.Its result has obtained 28 independently positive plaques.From wherein selecting 11 strong plaques of signal to carry out programmed screening.Programmed screening except that the second time washing time be according to carrying out 60 minutes with the same method of above-mentioned first time of screening.Its result has obtained the independent positive plaque that is produced by single phage from 6 clones.To insert the size of dna fragmentation part in order checking, as introduction, to carry out PCR as template with phage with above-mentioned R introduction and M13PrimerM4 (GTTTTCCCAGTCACGAC, precious wine is made) and M13PrimerRV (CAGGAAACAGCTATGAC, precious wine is made).As a result, 2 clones being arranged is total lengths.Then, carry out the live body cutting, in plasmid vector pBruescript II SK (-) (Stratagene company), make and insert dna fragmentation part inferior cloneization.The recombinant plasmid called after p411 and the p631 of inferior cloneization.

Find that by above-mentioned PCR the storehouse of Zhi Zuoing is the storehouse of containing the sufficiently long inset, being suitable for the cDNA screening as mentioned above.In a single day therefore,, advantageously adopt aforesaid method, and carry out aforesaid a large amount of RNA and handle, just can obtain a large amount of mRNA in order to separate the mRNA of pale brown colored chrysanthemum.

In addition, because above-mentioned cDNA storehouse contains aforesaid sufficiently long inset in a large number, thereby, can easily carry out the screening of full-length cDNA by using near the introduction the target protein working position as mentioned above to prepare the probe.

(2) comparison of determining and envision aminoacid sequence of the full base sequence of cDNA

In order to determine the segmental full base sequence of insertion cDNA of p631, made deletion mutant (Deletion mutants).The making of deletion mutant is carried out with the special-purpose Deletion Kit of Kilo-Sequence (precious wine is made company).Yet the reaction of exonuclease III is to make it to stop in 65 ℃ mung-bean nuclease buffer reagent by transferring to prior insulation.The determining to adopt with Qiagen Plasmid Mini Kit (Diagen company) purified plasmid, carry out of base sequence with Taq DyeDeoxy Terminator Cycle Sequencing Kit (ABI company) and Applied Biosystems 373A DNA Sequencer (ABI company).Article two, chain has all been determined its base sequence except that a part.According to determined base sequence, also determined aminoacid sequence.Determined base sequence and aminoacid sequence are illustrated in the sequence table sequence number 5.

It should be noted that, preparation be used for determining when separating the disappearance clone of base sequence of cDNA, importantly will be heated to 65 ℃ to the mung-bean nuclease damping fluid in advance, be that the disappearance reaction that exonuclease III is caused stops because just reaction solution being transferred in the mung bean nuclease damping fluid.In addition, because can't can be up to about 600～900bp base sequence partly from counting with the point of connection of carrier by only surveying from the upstream that gene that disappearance inserted determines, thereby be necessary to lack from both direction.

On the other hand, directly made with extra care PPDK, and determined the aminoacid sequence of its N-terminal zone, C-terminal zone and interior region from pale brown colored chrysanthemum.It is as follows that PPDK refining carries out.Grind with the extraction buffer reagent of 3 times of amounts and to add ammonium sulfate in the greenery of pale brown colored chrysanthemum → centrifugal back supernatant liquor and make it to reach 30% saturated and remove sedimentary protein → further add ammonium sulfate and make it to reach 70% saturated, reclaim albumen, the active fraction of the protein that adsorbs in the KCl concentration gradient drip washing post with Sephadex G25 (Pharmacia corporate system) desalination → → be associated with by DEASE-Sepharose post (Pharmacia corporate system) → with 50-400mM, concentrate with 70% saturated ammonium sulphate, with Sephadex G25 desalination → injection hydroxyapatite column → with the phosphoric acid concentration gradient is the active fraction of the protein that adsorbs on the phosphoric acid buffer agent drip washing post of 10mM～40mM → be associated with, concentrate with 70% saturated ammonium sulphate, with Sephadex G25 desalination.After this carry out the SDS-PAGE electrophoresis, cut the PPDK band, the electricity consumption stripping method reclaims the protein in the gel.Refining with this technology, obtain the about 5～10nmol of PPDK sample.The refining PPDK that obtains like this shows single band on SDS-PAGE.

Then, N-terminal sequence, C-terminal sequence and the internal sequence of resulting refining PPDK have been determined.That is, the aminoacid sequence of N-terminal is by transcribing protein on pvdf membrane, determining with the amino acid cut preface of gas phase device.The C-terminal sequence is with carboxypeptidase y digestion purified PPDK, infers according to the amino acid composition that dissociates out and the relation of digestion time.Inner aminoacid sequence is to distinguish what aminoacid sequence was determined by the N-terminal of the peptide that produces from protease digestion protein the time.Detailed method is as follows.At first, the method when determining the N-terminal aminoacid sequence is refined to the PPDK that the greenery from pale brown colored chrysanthemum obtain to a certain degree, carries out after the common SDS-PAGE electrophoresis, and gel cuts the PPDK band with Coomassie Brilliant BlueR250 dyeing.The gel that is cut in equilibration and the intervening well, carries out the SDS-PAGE electrophoresis second time in equilibration buffer reagent (Tris-HCl pH 6.8 125mM, EDTA 1mM, 0.1%SDS).At this moment, with the gel of equilibration with double-layer liquid (Tris-HCl pH6.8 125mM, EDTA 1mM, 0.1% SDS, 0.01% BPB, 20% glycerine) and enzyme solution (Tris-HCl pH6.8 125mM, EDTA 1mM, 0.1%SDS, 0.01%BPB, 10% glycerine, lysyl peptide ending enzyme 1～5 μ g or V8 proteinase-10 .01～0.1 μ g) add wait a moment electrophoresis after, in concentrated gel, carry out protein digestion (cut off the electricity supply, place 45 minutes).After this open electrophoresis again, the method when determining the N-terminal aminoacid sequence is transcribed on the pvdf membrane, determines the aminoacid sequence of digestion fragment from N-terminal with gas phase amino acid cut preface device.

The N-terminal sequence of Que Dinging, C-terminal sequence and internal sequence are as follows as mentioned above.

N-terminal sequence: Asn Pro Val Ser Pro Pro Val (72～78)

C-terminal sequence: Leu-Ala Ala *-Val Val (948～955)

Internal sequence (1): Lys Leu Tyr Gly Glu Phe Leu Val Asn Ala Gln Gly-Asp ValVal Ala (349～365)

Internal sequence (2): Gln Leu Leu Ala Pro Pro Ala Met Ser Asn Ala Leu-Thr (592～605)

Internal sequence (3): Leu Thr Ala Asp Thr Gly Met Ser Lys Asp Glu Ile Tyr Ser ArgIle Glu (721～738)

Internal sequence (4): Ala---Ser Phe Gly Thr Asn Asp Leu Cys Gln Met ValPhe Gly-Ser (844～862)

In addition, in above-mentioned aminoacid sequence, " * " means glutamine but can not be with used analytical instrument analysis, and "-" means that analytical results is unclear.Number in the bracket means the amino acid number of aminoacid sequence corresponding part shown in the sequence number 5.Some is different with the sequence of sequence number 5 corresponding parts in internal sequence (2) and (4), but this can think due to the error of amino acid sequencer.As everyone knows, the error of amino acid sequencer takes place with quite high frequency, and the error of DNA sequencer then takes place hardly.

Aminoacid sequence shown in the sequence number 5 has good consistence with the partial amino-acid series of directly determining from above-mentioned refining PPDK, thereby can confirm that aminoacid sequence shown in the sequence number is exactly the aminoacid sequence of PPDK.In addition, when the aminoacid sequence of aminoacid sequence shown in the sequence number 5 and well-known bidentate chrysanthemum chrysanthemum PPDK and corn PPDK compares, find that the different amino acid of mature protein part is respectively 40 (bidentate chrysanthemum chrysanthemums) and about 180 (corns).Can also recognize that from The above results No. 1 to No. 71 aminoacid sequence does not exist in aminoacid sequence shown in the sequence number 5 in mature protein, but a kind of during by film necessity pass on peptide, forming by the film post-treatment.The difference of amino-acid residue as shown in the following Table 1 in the mature protein of pale brown colored chrysanthemum and bidentate chrysanthemum chrysanthemum.

Table 1

	Pale brown colored chrysanthemum		Yellow tooth chrysanthemum chrysanthemum
				Different amino acid number and amino acid	5 Phe 10 Pro 15 Asn 28 Thr 40 Pro 41 Ala 48 Arg 49 Arg 50 Lys 52 Ser 57 Ile 61 Thr 62 Gly 66 Leu 81 Thr 92 Asn 97 Lys 114 Ala 140 Lys 150 Leu 154 Gln		6 Ser 11 Leu 16 Arg 29 Asn 41 Ser 42 Ser 47 Leu 48 Thr 49 Pro 50 Ala 55 Pro 59 Ser 60 Ser 64 Pro 79 Ala 90 Arg 95 Arg 112 Ser 138 Ser 148 Ser 152 Asp

170 Ala 201 Asp 205 Ala 263 Ala 265 Gln 323 Cys 356 Val 374 Val 382 Glu 385 Arg 392 Val 396 Arg 465 Asn 490 Val 564 Val 583 Ser 605 Thr 672 Ala 723 Thr 724 Ala 730 Lys 736 Arg 739 Lys 777 Asn 803 Gly 818 Leu 838 Asp 841 Ala

168 Pro 199 Ala 203 Gly 261 Val 263 Lys 321 Ser 354 Ile 372 Gly 380 Asp 383 Lys 390 Glu 394 Gly 463 Asp 488 Cys 562 Ile 581 Thr 603 Ile 670 Val 721 Ala 722 Val 728 Ala 734 Lys 737 Asn 775 Thr 801 Ser 816 Val 836 Glu 839 Gly

845 Glu 871 Pro 875 Ser 887 Leu 954 Val

843 Asp 869 Gln 873 Ala 885 Ile 952 Ile

2. the production of pale brown colored chrysanthemum PPDK in colibacillus and the mensuration of lower temperature resistance thereof

In order to confirm to have produced the lower temperature resistance enzyme really from isolating pale brown colored chrysanthemum PPDK cDNA as mentioned above, the expression in colibacillus is carried out as follows.

Combine in order to remove to pass on peptide and make expression vector and understand password, the importing at restriction enzyme position is carried out as follows.Cut off p631Sac (having lacked the p631 plasmid of SacI downstream area) that cyclisation again obtains as template with p631 with SacI, in order near the sequence the working position for the basis, contain the introduction 4:GATATCAATCCGGTGTCTCCTCC of EcoRV and carrier sequence complementary introduction M13RV (precious wine is made) be combined into performing PCR, and make the fragment of expansion in pCR II, carry out inferior cloneization.With restriction enzyme EcoRV and SacI, the fragment that contains the N-terminal part is scaled off from pCR II, cut off, make the pKK233-2 that cuts off with Hind III after the terminal smoothing carry out three segmental ligations (Fig. 1) together with the SacI-Hind III (nubbin that contains PPDK CDNA) of p631 with NcoI again with the Klenow enzyme.Make E.coli MV1184 transition and be used for expressing experiment with this plasmid.Nutrient solution adds IPTG 37 ℃ of shaking culture and makes it to reach 5mM with the new LB substratum of 9ml (containing the 50mg/l penbritin) dilution before the 1ml after 3 hours, further cultivates after 3 hours and uses the reclaiming by centrifuge thalline.Thalline is suspended in 0.5ml extraction buffer reagent (50mMHepes-KOH pH 7.5,10mM MgSO ₄, 1mM EDTA, 5mM DTT) in, add N,O-Diacetylmuramidase and make it to reach about 0.5mg/ml, handle 5 minutes on ice after, with ultrasonic disruption device (COSMOBIO UCD-130T type) limit with ice-cooled limit by handling the extraction enzyme 30 seconds at interval 5 minutes.With Eppendorf centrifuge centrifugal 10 minutes, supernatant liquor was by with post buffer reagent (50mM Hepes-KOH pH7.0,10mM MgCl ₂, 2mM EDTA after 10mMDTT) the Sephadex G25 post of equilibration is removed lower-molecular substance, placed 30 minutes at 25 ℃, carried out 4 aggressiveness and associated, and was used for determination of activity.

The PPDK that produces from the PPDK cDNA of pale brown colored chrysanthemum, bidentate chrysanthemum chrysanthemum and corn (Harvest Queen) in colibacillus demonstrates and comes from the enzyme mobility much at one of plant at SDS-PAGE.All roughly the same from the molecular weight of the maturing enzyme of various cDNA expection, but the apparent molecular weight on SDS-PAGE then has quite big-difference.This obviously is because the amino acid that is closed in each polypeptide is formed difference, rather than because the posttranslational modification of processing of proteinic disappearance or sugar chain addition etc.The lower temperature resistance of the various PPDK that produce with colibacillus is consistent with corresponding plant enzyme.That is, the lower temperature resistance of pale brown colored chrysanthemum PPDK need not just can obtain with the processing after this plant specific cofactor or the translation, if being expressed in this cDNA importing corn, just be expected to produce lower temperature resistance PPDK.Fig. 2 shows the relation of enzyme between the relative reactivity of time of being experienced after 0 ℃ the temperature placement and PPDK.

In addition, in above-mentioned technology, in order to produce activated PPDK with colibacillus, make removed pass on peptide-coding region cDNA and be inserted in the expression vector, but importantly make the location matches that will cut in the position corresponding with the N-terminal position of the enzyme that comes from plant this moment exactly.

(pale brown colored chrysanthemum) output (employed introduction) in colibacillus

MITAKKRVFTF… - 2

MIPVSPPVTTTKKRVFTF… + 3

MINPVSPPVTTTKKRVFTF… ++ 4

NPVSPPVTTTKKRVFTF ... (coming from the enzyme of leaf)

(bidentate chrysanthemum chrysanthemum)

MITAKKRVFTF… ++

MIPVSPPVTTAKKRVFTF… -

TAKKRVFTF ... (coming from the enzyme of leaf)

(corn)

MATKKRVFTF… ++

TTKKRVFTF ... (coming from the enzyme of leaf)

That is, when the expression experiment of carrying out pale brown colored chrysanthemum cDNA, the both successful face of land has reached corn and bidentate chrysanthemum chrysanthemum, also can express with reference to the place of incision of corn.Therefore,, use introduction 2 to cut off to make expression vector and attempt yet and expressed, but PPDK does not produce at all in same position even in pale brown colored chrysanthemum cDNA.Then,, made the expression vector that contains 7 residues at N-terminal more, when attempting to be expressed, confirmed to have produced PPDK with introduction 3 based on the N-terminal sequence of the enzyme that comes from leaf.But,, can't obtain being enough to measure the enzyme amount of lower temperature resistance even also seldom at this stage expression amount.Then, further made at N-terminal and surveyed the expression vector that prolongs 1 residue, when expressing experiment, can produce a large amount of PPDK, thereby can confirm its lower temperature resistance of introduction 4.The base sequence of introduction 2 and introduction 3 is as follows:

Introduction-2:CGGTGTCTCCTCCGGATATCACGGCTAAAAAGAG

Introduction-3:TTGATATCCCGGTTGTCTCCTCCGGTA

3. give determining of lower temperature resistance zone in the pale brown colored chrysanthemum PPDK gene

(1) mosaic of pale brown colored chrysanthemum and bidentate chrysanthemum chrysanthemum

Recombinate with restriction enzyme by well-established law.The homologous segment of the EcoRI-Hind III fragment that makes pKK-brownii and pKK-bidentis (adopting the method identical with the construction process of pKK-brownii the cDNA of bidentate chrysanthemum chrysanthemum to be inserted among the pKK-223-2 and the plasmid that obtains) exchanges makes pKK-011, otherwise, the homologous segment of the EcoR I-Hind III fragment of pKK-bidentis and pKK-brownii is exchanged makes pKK-100.Equally, NdeI-Hind III fragment is exchanged mutually and made pKK-001 and pKK-110.In addition, make the Xho-Hind III fragment of pKK-110 and the homologous segment exchange of pKK-bidentis make pKK-1101, and make the XhoI-Hind III fragment of pKK-bidentis and the homologous segment exchange of pKK-brownii make pKK-1110.In order to make the further reorganization mutually subtly of XhoI-Hind III fragment, carry out these segmental connections with PCR method (linking PCR method).Under the situation that base sequence is identical in the XhoI-Hind III fragment of bidentis (bidentate chrysanthemum chrysanthemum) and brownii (pale brown colored chrysanthemum), complementary introduction link-F:GCAGAGATGATGTTGGCAAG and link-R:CTTGCCAACATCATCTCTGC have been made.With with the XhoI-Hind III fragment of the brownii of pBluescript SK (-) inferior cloneization or bidentis as template, carry out the PCR first time with the combination of link-F/RV, M4/link-R.It is refining that resulting fragment (totally 4 kinds) cuts out method with gel, as template, carries out second time PCR with introduction M4 RV with brownii forebody and bidentis is latter half of or bidentis forebody and brownii are latter half of fragment mixture.The binding fragment that enlarges cuts off with XhoI and Hind III, with the corresponding part exchange of pKK-bidentis, has made pKK-link01 and pKK-link10.Utilize the reorganization position of linking PCR and the PstI position between the Hind III, made another group chimeric gene.XhoI-Hind III position (3 fragment ligation) the PstI-Hind III fragment of the XhoI-PstI fragment of pKK-link10 and pKK-bidentis is inserted pKK-bidentis has made pKK-link101.The XhoI-PstI fragment of pKK-bidentis and the PstI-HindIII fragment of pKK-brownii also by the XhoI-HindIII position of inserting pKK-bidentis with quadrat method, have been made pKK-link110.

The PPDK of pale brown colored chrysanthemum and bidentate chrysanthemum chrysanthemum at the amino-acid substitution at 40 positions that mature protein partly occurs morely near the N-terminal and the C-terminal zone of enzyme, and less near the active centre, middle part.Therefore, at first utilize in two kinds of genes common restriction enzyme EcoRI that exists and NdeI position that cDNA is divided into before, during and after three parts, these fragments are exchanged mutually make chimeric gene, investigated which part is relevant with lower temperature resistance on earth.Consequently, after containing pale brown colored chrysanthemum cDNA, obtained lower temperature resistance under 1/3 the situation, otherwise, after containing bidentate chrysanthemum chrysanthemum cDNA, have low-temperature sensitive under 1/3 the situation.Then, 1/3 zone, back is divided into two parts, imports respectively in the corresponding part of pKK-bidentis, measure lower temperature resistance with restriction enzyme XhoI.Consequently, the necessity and the sufficient condition of acquisition lower temperature resistance are the downstream areas (apart from the zone of C least significant end 1/6) that contains the XhoI position.Then, make the chimeric gene that contains 1/6 zone, back (XhoI-Hind III fragment contains 7 amino-acid substitutions), import among the pKK-bidentis, measure its lower temperature resistance with linking PCR method.Consequently, have the mosaic enzyme pKK-link10 that contains 4 metathetical final areas lower temperature resistance arranged, have contain half zone before 3 metathetical mosaic enzyme pKK-link01 then at low-temperature inactive.Therefore, make this later half zone reorganization with restriction enzyme PstI, made the chimeric gene that 2 amino-acid substitutions are respectively arranged, measure lower temperature resistance, but wherein any one mosaic enzyme all shows lower temperature resistance, thereby there is zone relevant with lower temperature resistance more than 2 in supposition.

(2) mosaic of corn and pale brown colored chrysanthemum

With introduction PPDK-F:CTCACTGTTCGAAGAGAAGC and the mNdeI:CATATGCTCTGTCCGGCATAATC (on the complementary strand) that contains the NdeI position, be that template is carried out PCR with corn PPDK cDNA, resulting fragment carry out inferior cloneization in pCR II.With SacI and NdeI this fragment is scaled off from pCR II, the ScaI-SmaI fragment (carrier segments) of pKK-PPDK and pale brown colored chrysanthemum PPDK cDNA cut off with Hind III, make after the terminal smoothing with the Klenow enzyme and to cut off the fragment that obtains with NdeI, it is carried out 3 fragment ligations (Fig. 3) together, obtained pKK-mz/bro (Nde).In addition, is that template carry out PCR with the mXhoI:CTCGAGGGATCTCAATCATTG (on the complementary strand) that contains the XhoI position with corn PPDK cDNA with introduction PPDK-F, resulting fragment scales off from pCR II with SacI and Xho I after the inferior cloneization in pCR II, combine with the ScaI-XhoI fragment (carrier segments) of pKK-mz/bro (Nde), obtain pKK-mz/bro (Xho).

The mosaic enzyme of corn PPDK and pale brown colored chrysanthemum PPDK C-terminal 1/3 zone (NdeI-Hind III fragment) or 1/6 zone (XhoI-Hind III fragment) all shows the strong lower temperature resistance with pale brown colored chrysanthemum par.Like this, just can make the rather different corn PPDK of aminoacid sequence have lower temperature resistance, thereby can think, just can make the PPDK that comes from each kind of plant have lower temperature resistance by importing this last 1/3 zone or 1/6 zone.Because the corn/pale brown colored chrysanthemum mosaic PPDK that makes can be used with its original form to the peptide that passes on of corn PPDK thus, thereby, then can consider to solve with the way that this chimeric gene of importing replaces if it passes on body transition that part also comes from pale brown colored chrysanthemum PPDK and on chloroplast(id) is carried problem is arranged at lower temperature resistance PPDK.

(3) variation clone

For the XhoI-Hind III fragment of pKK-brownii, made the amino-acid residue of pale brown colored chrysanthemum → bidentate chrysanthemum chrysanthemum type and replaced one by one.And, also made the amino-acid residue of bidentate chrysanthemum chrysanthemum type → pale brown colored chrysanthemum type and replaced one by one for the XhoI-Hind III fragment of pKK-bidentis.It is XhoI-Hind III fragment inferior cloneization in pBluescript II SK (-) respectively by making pale brown colored chrysanthemum and bidentate chrysanthemum chrysanthemum PPDK cDNA that variation imports, and uses Mega label Kit (precious wine is made) and Mutan-K Kit (precious wine is made) to carry out according to the Kunkel method again.The introduction sequence that imports of being used to make a variation is as shown in table 2.The base sequence of variation inserts these fragments in the XhoI-Hind III position of pKK-bidentis after confirming with the DNA sequencer.

Table 2 is used for an introduction that the variation clone is made

The variation of pale brown colored chrysanthemum type → bidentate chrysanthemum chrysanthemum type imports
	836DE
5′GCAATCTCTTCAGCAATC 839AG 5′GCTTCTTTTCCAATCTCATC 843ED 5′CGAAAAGAAATCGGCTTC 869PQ 5′GAAAGATAAATCTGCAAAAACTTG 873SA 5′GCCTTGAGCAAGATAAATC 885LI 5′TTCTGGTCAATAACCTCAATG 952VI 5′GCTTAAACAATGACTTGTGC
	The variation of bidentate chrysanthemum chrysanthemum type → pale brown colored chrysanthemum type imports
836ED
	5′CCAATCTCATCAGCTATTAAAG 839GA 5′GCTTCTTTTGCAATCTCTTC 843DE 5′CGAAAAGAACTCAGCTTC 869QP 5′CAAGATAAATCGGCAAAAACTTG 873AS 5′GAATGCCTTGAGAAAGATAAATC 885IL 5′CTTTCTGGTCAAGAACCTCAAATG 952IV 5′GCTTAAACAACGACTTGTGC

The enzyme that makes different amino acid in the XhoI-Hind III zone of pKK-1110 be replaced as bidentate chrysanthemum chrysanthemum type one by one all shows lower temperature resistance, thereby can think and exist a plurality of variations (only changing a residue without detriment to resistance) of giving lower temperature resistance.Then, in contrast, make that different amino acid is replaced as pale brown colored chrysanthemum type one by one in the XhoI-Hind III zone of pKK-bidentis, investigate it and whether become the lower temperature resistance enzyme.That is, after 20 minutes, measure enzymic activity 0 ℃ of processing.Consequently, 896Gln → Pro variation obtains lower temperature resistance (activity after the above-mentioned subzero treatment be original 60～70%), and 885Ile → Leu and 952Ile → Val variation makes that low-temperature inactive is difficult to take place slightly, if take all factors into consideration the result of above-mentioned (1) mosaic enzyme pKK-link110, still can obtain lower temperature resistance in the time of then can inferring these two kinds variation coexistences.It can be asserted that 869Pro, 885Leu, these three residues of 925Val are relevant with lower temperature resistance from above result.Yet in pale brown colored chrysanthemum these residues relevant with lower temperature resistance, with regard to 869Pro and 885Leu, corn PPDK also has pale brown colored chrysanthemum type.Therefore, only these residues be pale brown colored chrysanthemum type not necessarily just can obtain lower temperature resistance, but can think that they have just given resistance completely at first in the aminoacid sequence of pale brown colored chrysanthemum or bidentate chrysanthemum chrysanthemum, thereby, under the situation that makes quite big other species PPDK lower temperature resistanceization of aminoacid sequence difference, should not import a variation, better be as corn PPDK being carried out, making the chimeric gene that imports with area unit.

4. the corn that carries out with pale brown colored chrysanthemum PPDK gene makes the transition

According to people (The Plant Cell 2:603-618 such as Gordon-Kamm W.J., 1990) or people (Bio/Technology 11:194-200 such as Koziel M.G., 1993) method, pKK-brownii is coated on the minuteness particle of tungsten or gold, it is implanted in the immature embryo or suspended culture cell of corn.From importing the cell screening cell of handling transition, then, resulting body callus transition (callus) is pressed well-established law cultivate the regeneration value object.Transition, method was not limited to the particle marksmanship, also comprise Electro-poration method (Rhodes C.A.et al., Science 240:204-207,1988), PEG method (Armstrong C.L.et al., Plant Cell Reports 9:335-339,1990), Tissue-electroporation method (D ' Halluin K.et al., ThePlant Cell 4:1495-1505,1992) or edaphic bacillus method (Hiei Y.andKomari T., WO 9400977) etc.Reclaim seed and make it germination from resulting plant materials, the leaf from resulting plant separates PPDK, measures its lower temperature resistance again.Measure temperature to transformed plant and the not influence of transformed plant light compositing speed.By making those show that at low temperature milpas of high light resultant velocity breed many generations, measure the light compositing speed of differing temps and measure lower temperature resistance from the isolating PPDK of plant materials, guaranteed the stability of transition of being studied.

5. the bidentate chrysanthemum chrysanthemum that carries out with pale brown colored chrysanthemum PPDK gene makes the transition

The intermediate carrier that contains full-length cDNA shown in the sequence number 5 and contain information gene import the swelling edaphic bacillus ( Agrobacterium Tumefaciens) innoxious Ti-plasmids in.This can be according to Draper J et al.eds., Plant Genetic Transfomationand Gene Expression-a laboratory manual, method is carried out described in the BlackwellScientific Publications (ISBN 0-632-02172-1).

On the other hand, make leaf texture of bidentate chrysanthemum chrysanthemum (F.bidentis) or callus (callus) infect above-mentioned swelling edaphic bacillus (A.tumefaciens).This can be undertaken by the coexistence of this tissue or callus and swelling edaphic bacillus is cultivated.Cells infected is selected according to resistance.Press well-established law aftergrowth body from selected callus.Obtain seed, make it to germinate from resultant plant materials, separate PPDK and measure its lower temperature resistance from the leaf of resulting plant.For transformed plant and transformed plant not, investigate the influence of temperature to light compositing speed.By making those show that at low temperature bidentate chrysanthemum chrysanthemum plant of high light resultant velocity breed many generations, measure the light compositing speed of differing temps and measure lower temperature resistance from the isolating PPDK of plant materials, guarantee the stability of transition of being studied.

[sequence table]

Sequence number: 1

Sequence is long: 2915

Sequence type: nucleic acid

Sequence

CGATCTCCTT CTGCTATTGC TGATATCTCA ATTTCACAGG TGAAGAAGG ATG ATG AGT 58

Met Met Ser

1

TCG TTG TCT GTT GAA GGT ATG CTT CTC AAG TCA GCC CGT GAG TCG TGC 106

Ser Leu Ser Val Glu Gly Met Leu Leu Lys Ser Ala Arg Glu Ser Cys

5 10 15

TTA CCG GCG AGA GTG AAG CAA CGG CGA AAC GGT GAT CTC CGG CGA TTG 154

Leu Pro Ala Arg Val Lys Gln Arg Arg Asn Gly Asp Leu Arg Arg Leu

20 25 30 35

AAC CAC CAC CGT CAA TCG TCG TTT GTC CGG TGT TTA ACT CCG GCG AGA 202

Asn His His Arg Gln Ser Ser Phe Val Arg Cys Leu Thr Pro Ala Arg

40 45 50

GTT AGC AGA CCA GAG TTG CGC AGC AGT GGC TTA ACT CCG CCG CGA GCA 250

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

55 60 65

GTT CTT AAT CCG GTG TCT CCT CCG GTG ACG ACG GCT AAA AAG AGG GTT 298

Val Leu Asn Pro Val Ser Pro Pro Val Thr Thr Ala Lys Lys Arg Val

70 75 80

TTC ACT TTT GGT AAA GGA AGA AGT GAA GGC AAC AGG GAC ATG AAA TCC 346

Phe Thr Phe Gly Lys Gly Arg Ser Glu Gly Asn Arg Asp Met Lys Ser

85 90 95

TTG TTG GGA GGA AAA GGA GCA AAT CTT GCT GAG ATG TCA AGC ATT GGT 394

Leu Leu Gly Gly Lys Gly Ala Asn Leu Ala Glu Met Ser Ser Ile Gly

100 105 110 115

CTA TCA GTT CCT CCT GGG CTC ACT ATT TCA ACT GAA GCA TGT GAG GAA 442

Leu Ser Val Pro Pro Gly Leu Thr Ile Ser Thr Glu Ala Cys Glu Glu

120 125 130

TAT CAA CAA AAT GGA AAG AGC CTA CCT CCA GGT TTG TGG GAT GAG ATT 490

Tyr Gln Gln Asn Gly Lys Ser Leu Pro Pro Gly Leu Trp Asp Glu Ile

135 140 145

TCA GAA GGC TTA GAT TAT GTC CAG AAA GAG ATG TCT GCA TCT CTC GGT 538

Ser Glu Gly Leu Asp Tyr Val Gln Lys Glu Met Ser Ala Ser Leu Gly

150 155 160

GAC CCG TCT AAA CCT CTC CTC CTT TCC GTC CGT TCG GGT GCT GCC ATA 586

Asp Pro Ser Lys Pro Leu Leu Leu Ser Val Arg Ser Gly Ala Ala Ile

165 170 175

TCT ATG CCT GGT ATG ATG GAC ACT GTA TTG AAT CTC GGG CTT AAT GAT 634

Ser Met Pro Gly Met Met Asp Thr Val Leu Asn Leu Gly Leu Asn Asp

180 185 190 195

GAG GTC GTA GCT GGT CTA GCT GGC AAA AGT GGA GCA CGG TTT GCC TAT 682

Glu Val Val Ala Gly Leu Ala Gly Lys Ser Gly Ala Arg Phe Ala Tyr

200 205 210

GAC TCG TAT AGA AGG TTT CTC GAT ATG TTT GGC AAC GTT GTA ATG GGT 730

Asp Ser Tyr Arg Arg Phe Leu Asp Met Phe Gly Asn Val Val Met Gly

215 220 225

ATC CCG CAT TCA TTA TTT GAC GAA AAG TTA GAG CAG ATG AAA GCT GAA 778

Ile Pro His Ser Leu Phe Asp Glu Lys Leu Glu Gln Met Lys Ala Glu

230 235 240

AAA GGG ATT CAT CTC GAC ACC GAT CTC ACT GCT GCT GAT CTT AAA GAT 826

Lys Gly Ile His Leu Asp Thr Asp Leu Thr Ala Ala Asp Leu Lys Asp

245 250 255

CTT GTT GAG AAA TAC AAG AAC GTG TAT GTG GAA GCA AAG GGC GAA AAG 874

Leu Val Glu Lys Tyr Lys Asn Val Tyr Val Glu Ala Lys Gly Glu Lys

260 265 270 275

TTT CCC ACA GAT CCA AAG AAA CAG CTA GAG TTA GCA GTG AAT GCT GTT 922

Phe Pro Thr Asp Pro Lys Lys Gln Leu Glu Leu Ala Val Asn Ala Val

280 285 290

TTT GAT TCT TGG GAC AGC CCA AGG GCC AAT AAG TAC AGA AGT ATT AAC 970

Phe Asp Ser Trp Asp Ser Pro Arg Ala Asn Lys Tyr Arg Ser Ile Asn

295 300 305

CAG ATA ACT GGA TTA AAG GGG ACT GCA GTT AAC ATT CAA AGC ATG GTG 1018

Gln Ile Thr Gly Leu Lys Gly Thr Ala Val Asn Ile Gln Ser Met Val

310 315 320

TTT GGC AAC ATG GGA AAC ACT TCA GGA ACT GGT GTT CTT TTC ACT AGG 1066

Phe Gly Asn Met Gly Asn Thr Ser Gly Thr Gly Val Leu Phe Thr Arg

325 330 335

AAC CCA AGC ACC GGT GAG AAG AAG CTA TAC GGG GAG TTT TTA ATC AAT 1114

Asn Pro Ser Thr Gly Glu Lys Lys Leu Tyr Gly Glu Phe Leu Ile Asn

340 345 350 355

GCT CAG GGA GAG GAT GTT GTT GCT GGG ATC AGA ACA CCA GAA GAT TTG 1162

Ala Gln Gly Glu Asp Val Val Ala Gly Ile Arg Thr Pro Glu Asp Leu

360 365 370

GGG ACC ATG GAG ACT TGC ATG CCT GAT GCA TAC AAA GAG CTT GTG GAG 1210

Gly Thr Met Glu Thr Cys Met Pro Asp Ala Tyr Lys Glu Leu Val Glu

375 380 385

AAC TGC GAG ATC TTA GAG GGA CAC TAC AAA GAT ATG ATG GAT ATT GAA 1258

Asn Cys Glu Ile Leu Glu Gly His Tyr Lys Asp Met Met Asp Ile Glu

390 395 400

TTC ACA GTT CAA GAA AAC AGG CTT TGG ATG TTG CAA TGC CGA ACA GGG 1306

Phe Thr Val Gln Glu Asn Arg Leu Trp Met Leu Gln Cys Arg Thr Gly

405 410 415

AAA CGT ACT GGT AAA GGT GCA GTG AGA ATT GCA GTA GAT ATG GTG AAC 1354

Lys Arg Thr Gly Lys Gly Ala Val Arg Ile Ala Val Asp Met Val Asn

420 425 430 435

GAA GGG CTA ATT GAT ACT AGA ACA GCA ATT AAG AGG GTT GAG ACT CAA 1402

Glu Gly Leu Ile Asp Thr Arg Thr Ala Ile Lys Arg Val Glu Thr Gln

440 445 450

CAT CTA GAT CAG CTT CTT CAT CCA CAG TTT GAG GAT CCG TCT GCT TAC 1450

His Leu Asp Gln Leu Leu His Pro Gln Phe Glu Asp Pro Ser Ala Tyr

455 460 465

AAA AGC CAT GTG GTA GCA ACC GGT TTG CCA GCA TCC CCC GGG GCA GCT 1498

Lys Ser His Val Val Ala Thr Gly Leu Pro Ala Ser Pro Gly Ala Ala

470 475 480

GTG GGA CAG GTT TGT TTT AGT GCA GAG GAT GCA GAA ACA TGG CAT GCA 1546

Val Gly Gln Val Cys Phe Ser Ala Glu Asp Ala Glu Thr Trp His Ala

485 490 495

CAA GGA AAG AGT GCT ATC TTG GTA AGG ACC GAA ACA AGC CCA GAA GAT 1594

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

500 505 510 515

GTT GGT GGT ATG CAT GCA GCA GCT GGA ATC TTA ACC GCT AGA GGA GGC 1642

Val Gly Gly Met His Ala Ala Ala Gly Ile Leu Thr Ala Arg Gly Gly

520 525 530

ATG ACA TCA CAT GCA GCG GTG GTG GCT CGC GGA TGG GGC AAA TGT TGT 1690

Met Thr Ser His Ala Ala Val Val Ala Arg Gly Trp Gly Lys Cys Cys

535 540 545

GTT TCC GGT TGT GCT GAT ATT CGT GTG AAC GAT GAT ATG AAG ATT TTT 1738

Val Ser Gly Cys Ala Asp Ile Arg Val Asn Asp Asp Met Lys Ile Phe

550 555 560

ACG ATT GGC GAC CGT GTG ATT AAA GAA GGC GAC TGG CTT TCT CTT AAT 1786

Thr Ile Gly Asp Arg Val Ile Lys Glu Gly Asp Trp Leu Ser Leu Asn

565 570 575

GGT ACA ACT GGT GAA GTC ATA TTG GGT AAA CAG CTA CTG GCT CCA CCT 1834

Gly Thr Thr Gly Glu Val Ile Leu Gly Lys Gln Leu Leu Ala Pro Pro

580 585 590 595

GCA ATG AGC AAT GAC TTA GAA ATA TTC ATG TCA TGG GCT GAT CAA GCA 1882

Ala Met Ser Asn Asp Leu Glu Ile Phe Met Ser Trp Ala Asp Gln Ala

600 605 610

AGG CGT CTC AAG GTT ATG GCA AAT GCA GAC ACA CCT AAT GAT GCA TTA 1930

Arg Arg Leu Lys Val Met Ala Asn Ala Asp Thr Pro Asn Asp Ala Leu

615 620 625

ACA GCC AGA AAC AAT GGT GCA CAA GGG ATC GGG CTC TGT AGA ACT GAA 1978

Thr Ala Arg Asn Asn Gly Ala Gln Gly Ile Gly Leu Cys Arg Thr Glu

630 635 640

CAT ATG TTT TTC GCT TCT GAT GAG AGG ATC AAA GCT GTA AGA AAG ATG 2026

His Met Phe Phe Ala Ser Asp Glu Arg Ile Lys Ala Val Arg Lys Met

645 650 655

ATC ATG GCG GTC ACT CCA GAA CAA AGA AAA GTG GCT CTA GAT CTC TTA 2074

Ile Met Ala Val Thr Pro Glu Gln Arg Lys Val Ala Leu Asp Leu Leu

660 665 670 675

CTC CCA TAC CAA AGA TCC GAT TTT GAG GGC ATT TTC CGA GCA ATG GAT 2122

Leu Pro Tyr Gln Arg Ser Asp Phe Glu Gly Ile Phe Arg Ala Met Asp

680 685 690

GGA CTT CCT GTA ACT ATC CGC CTT CTA GAC CCT CCA CTT CAT GAG TTT 2170

Gly Leu Pro Val Thr Ile Arg Leu Leu Asp Pro Pro Leu His Glu Phe

695 700 705

TTA CCC GAA GGT GAT CTA GAA CAC ATA GTG AAC GAA CTT GCA GTC GAC 2218

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

710 715 720

ACA GGC ATG AGT GCA GAT GAA ATC TAT TCA AAA ATC GAA AAT CTA TCT 2266

Thr Gly Met Ser Ala Asp Glu Ile Tyr Ser Lys Ile Glu Asn Leu Ser

725 730 735

GAA GTG AAC CCT ATG CTT GGT TTC CGT GGT TGC AGA TTA GGG ATT TCA 2314

Glu Val Asn Pro Met Leu Gly Phe Arg Gly Cys Arg Leu Gly Ile Ser

740 745 750 755

TAC CCC GAG CTA ACA GAA ATG CAA GTT CGT GCG ATC TTT CAA GCT GCA 2362

Tyr Pro Glu Leu Thr Glu Met Gln Val Arg Ala Ile Phe Gln Ala Ala

760 765 770

GTG TCT ATG ACC AAT CAG GGG GTG ACT GTA ATA CCA GAG ATC ATG GTT 2410

Val Ser Met Thr Asn Gln Gly Val Thr Val Ile Pro Glu Ile Met Val

775 780 785

CCG TTA GTG GGG ACA CCT CAG GAA TTA CGT CAT CAA ATC AGT GTA ATT 2458

Pro Leu Val Gly Thr Pro Gln Glu Leu Arg His Gln Ile Ser Val Ile

790 795 800

CGT GGA GTA GCT GCA AAT GTG TTT GCT GAA ATG GGG GTG ACA TTG GAA 2506

Arg Gly Val Ala Ala Asn Val Phe Ala Glu Met Gly Val Thr Leu Glu

805 810 815

TAT AAA GTG GGA ACG ATG ATT GAG ATT CCT CGA GCT GCT TTA ATA GCT 2554

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

820 825 830 835

GAA GAG ATT GGA AAA GAA GCT GAT TTC TTT TCG TTT GGA ACC AAT GAT 2602

Glu Glu Ile Gly Lys Glu Ala Asp Phe Phe Ser Phe Gly Thr Asn Asp

840 845 850

CTG ACC CAG ATG ACA TTT GGG TAC AGC AGA GAT GAT GTT GGC AAG TTT 2650

Leu Thr Gln Met Thr Phe Gly Tyr Ser Arg Asp Asp Val Gly Lys Phe

855 860 865

TTG CAG ATT TAT CTT GCT CAA GGC ATT CTG CAG CAT GAT CCA TTT GAG 2698

Leu Gln Ile Tyr Leu Ala Gln Gly Ile Leu Gln His Asp Pro Phe Glu

870 875 880

GTT ATT GAC CAG AAA GGG GTG GGT CAG TTG ATT AAG ATG GCT ACG GAG 2746

Val Ile Asp Gln Lys Gly Val Gly Gln Leu Ile Lys Met Ala Thr Glu

885 890 895

AAA GGT CGT GCA GCA AAT CCT AAC TTA AAG GTT GGG ATA TGT GGG GAG 2794

Lys Gly Arg Ala Ala Asn Pro Asn Leu Lys Val Gly Ile Cys Gly Glu

900 905 910 915

CAT GGT GGG GAG CCT TCT TCT GTT GCA TTT TTT GAT GGA GTT GGA CTA 2842

His Gly Gly Glu Pro Ser Ser Val Ala Phe Phe Asp Gly Val Gly Leu

920 925 930

GAT TAT GTG TCG TGC TCT CCA TTT AGG GTT CCT ATC GCA AGG TTG GCC 2890

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

935 940 945

GCT GCA CAA GTC ATT GTT TAAGCTT 2915

Ala Ala Gln Val Ile Val

950

Sequence number: 2

Sequence is long: 2880

Sequence type: nucleic acid

Sequence

CGGCGCAGTA GGGGATCGGA AGG ATG GCG GCA TCG GTT TCC AGG GCC ATC TGC 53

Met Ala Ala Ser Val Ser Arg Ala Ile Cys

1 5 10

GTA CAG AAG CCG GGC TCA AAA TGC ACC AGG GAC AGG GAA GCG ACC TCC 101

Val Gln Lys Pro Gly Ser Lys Cys Thr Arg Asp Arg Glu Ala Thr Ser

15 20 25

TTC GCC CGC CGA TCG GTC GCA GCG CCG AGG CCC CCG CAC GCC AAA GCC 149

Phe Ala Arg Arg Ser Val Ala Ala Pro Arg Pro Pro His Ala Lys Ala

30 35 40

CGC CGG CGT CAT CCG CTC CGA CTC CGG CGC GGG ACG GGG CCA CAT TGC 197

Arg Arg Arg His Pro Leu Arg Leu Arg Arg Gly Thr Gly Pro His Cys

45 50 55

TCG CCG CTG AGG GCC GTC GTT GAC GCC GCG CCG ATA CAG ACG ACC AAA 245

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

60 65 70

AAG AGG GTG TTC CAC TTC GGC AAG GGC AAG AGC GAG GGC AAC AAG ACC 293

Lys Arg Val Phe His Phe Gly Lys Gly Lys Ser Glu Gly Asn Lys Thr

75 80 85 90

ATG AAG GAA CTG CTG GGC GGC AAG GGC GCG AAC CTG GCG GAG ATG GCG 341

Met Lys Glu Leu Leu Gly Gly Lys Gly Ala Asn Leu Ala Glu Met Ala

95 100 105

AGC ATC GGG CTG TCG GTG CCG CCA GGG TTC ACG GTG TCG ACG GAG GCG 389

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

110 115 120

TGC CAG CAG TAC CAG GAC GCC GGG TGC GCC CTC CCC GCG GGG CTC TGG 437

Cys Gln Gln Tyr Gln Asp Ala Gly Cys Ala Leu Pro Ala Gly Leu Trp

125 130 135

GCC GAG ATC GTC GAC GGC CTG CAG TGG GTG GAG GAG TAC ATG GGC GCC 485

Ala Glu Ile Val Asp Gly Leu Gln Trp Val Glu Glu Tyr Met Gly Ala

140 145 150

ACC CTG GGC GAT CCG CAG CGC CCG CTC CTG CTC TCC GTC CGC TCC GGC 533

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

155 160 165 170

GCC GCC GTG TCC ATG CCC GGC ATG ATG GAC ACG GTG CTC AAC CTG GGG 581

Ala Ala Val Ser Met Pro Gly Met Met Asp Thr Val Leu Asn Leu Gly

175 180 185

CTC AAC GAC GAA GTG GCC GCC GGG CTG G CG GCC AAG AGC GGG GAG CGC 629

Leu Asn Asp Glu Val Ala Ala Gly Leu Ala Ala Lys Ser Gly Glu Arg

190 195 200

TTC GCC TAC GAC TCC TTC CGC CCC TTC CTC GAC ATG TTC GGC AAC GTC 677

Phe Ala Tyr Asp Ser Phe Arg Arg Phe Leu Asp Met Phe Gly Asn Val

205 210 215

GTC ATG GAC ATC CCC CGC TCA CTG TTC GAA GAG AAG CTT GAG CAC ATG 725

Val Met Asp Ile Pro Arg Ser Leu Phe Glu Glu Lys Leu Glu His Met

220 225 230

AAG GAA TCC AAG GGG CTG AAG AAC GAC ACC GAC CTC ACG GCC TCT GAC 773

Lys Glu Ser Lys Gly Leu Lys Asn Asp Thr Asp Leu Thr Ala Ser Asp

235 240 245 250

CTC AAA GAG CTC GTG GGT CAG TAC AAG GAG GTC TAC CTC TCA GCC AAG 821

Leu Lys Glu Leu Val Gly Gln Tyr Lys Glu Val Tyr Leu Ser Ala Lys

255 260 265

GGA GAG CCA TTC CCC TCA GAC CCC AAG AAG CAG CTG GAG CTA GCA GTG 869

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

270 275 280

CTG GCT GTG TTC AAC TCG TGG GAG AGC CCC AGG GCC AAG AAG TAC AGG 917

Leu Ala Val Phe Asn Ser Trp Glu Ser Pro Arg Ala Lys Lys Tyr Arg

285 290 295

AGC ATC AAC CAG ATC ACT GGC CTC AGG GGC ACC GCC GTG AAC GTG CAG 965

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

300 305 310

TGC ATG GTG TTC GGC AAC ATG GGG AAC ACT TCT GGC ACC GGC GTG CTC 1013

Cys Met Val Phe Gly Asn Met Gly Asn Thr Ser Gly Thr Gly Val Leu

315 320 325 330

TTC ACC AGG AAC CCC AAC ACC GGA GAG AAG AAG CTG TAT GGC GAG TTC 1061

Phe Thr Arg Asn Pro Asn Thr Gly Glu Lys Lys Leu Tyr Gly Glu Phe

335 340 345

CTG GTG AAC GCT CAG GGT GAG GAT GTG GTT GCC GGA ATA AGA ACC CCA 1109

Leu Val Asn Ala Gln Gly Glu Asp Val Val Ala Gly Ile Arg Thr Pro

350 355 360

GAG GAC CTT GAC GCC ATG AAG AAC CTC ATG CCA CAG GCC TAC GAC GAG 1157

Glu Asp Leu Asp Ala Met Lys Asn Leu Met Pro Gln Ala Tyr Asp Glu

365 370 375

CTT GTT GAG AAC TGC AAC ATC CTG GAG AGC CAC TAC AAG GAA ATG CAG 1205

Leu Val Glu Asn Cys Asn Ile Leu Glu Ser His Tyr Lys Glu Met Gln

380 385 390

GAT ATC GAG TTC ACT GTC CAG GAA AAC AGG CTG TGG ATG TTG CAG TGC 1253

Asp Ile Glu Phe Thr Val Gln Glu Asn Arg Leu Trp Met Leu Gln Cys

395 400 405 410

AGG ACA GGG AAA CGT ACG GGC AAA AGT GCC GTG AAG ATC GCC GTG GAC 1301

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

415 420 425

ATG GTT AAC GAG GGC CTT GTT GAG CCC CGC TCA GCG ATC AAG ATG GTA 1349

Met Val Asn Glu Gly Leu Val Glu Pro Arg Ser Ala Ile Lys Met Val

430 435 440

GAG CCA GGC CAC CTG GAC CAG CTT CTT CAT CCT CAG TTT GAG AAC CCG 1397

Glu Pro Gly His Leu Asp Gln Leu Leu His Pro Gln Phe Glu Asn Pro

445 450 455

TCG GCG TAC AAG GAT CAA GTC ATT GCC ACT GGT CTG CCA GCC TCA CCT 1445

Ser Ala Tyr Lys Asp Gln Val Ile Ala Thr Gly Leu Pro Ala Ser Pro

460 465 470

GGG GCT GCT GTG GGC CAG GTT GTG TTC ACT GCT GAA GAT GCT GAA GCA 1493

Gly Ala Ala Val Gly Gln Val Val Phe Thr Ala Glu Asp Ala Glu Ala

475 480 485 490

TGG CAT TCC CAA GGG AAA GCT GCT ATT CTG GTA AGG GCG GAG ACC AGC 1541

Trp His Ser Gln Gly Lys Ala Ala Ile Leu Val Arg Ala Glu Thr Ser

495 500 505

CCT GAG GAC GTT GGT GGC ATG CAC GCT GCT GTG GGG ATT CTT ACA GAG 1589

Pro Glu Asp Val Gly Gly Met His Ala Ala Val Gly Ile Leu Thr Glu

510 515 520

AGG GGT GGC ATG ACT TCC CAC GCT GCT GTG GTC GCA CGT TGG TGG GGG 1637

Arg Gly Gly Met Thr Ser His Ala Ala Val Val Ala Arg Trp Trp Gly

525 530 535

AAA TGC TGC GTC TCG GGA TGC TCA GGC ATT CGC GTA AAC GAT GCG GAG 1685

Lys Cys Cys Val Ser Gly Cys Ser Gly Ile Arg Val Asn Asp Ala Glu

540 545 550

AAG CTC GTG ACG ATC GGA AGC CAT GTG CTG CGC GAA GGT GAG TGG CTG 1733

Lys Leu Val Thr Ile Gly Ser His Val Leu Arg Glu Gly Glu Trp Leu

555 560 565 570

TCG CTG AAT GGG TCG ACT GGT GAG GTG ATC CTT GGG AAG CAG CCG CTT 1781

Ser Leu Asn Gly Ser Thr Gly Glu Val Ile Leu Gly Lys Gln Pro Leu

575 580 585

TCC CCA CCA GCC CTT AGT GGT GAT CTG GGA ACT TTC ATG GCC TGG GTG 1829

Ser Pro Pro Ala Leu Ser Gly Asp Leu Gly Thr Phe Met Ala Trp Val

590 595 600

GAT GAT GTT AGA AAG CTC AAG GTC CTG GCT AAC GCC GAT ACC CCT GAT 1877

Asp Asp Val Arg Lys Leu Lys Val Leu Ala Asn Ala Asp Thr Pro Asp

605 610 615

GAT GCA TTG ACT GCG CGA AAC AAT GGG GCA CAA GGA ATT GGA TTA TGC 1925

Asp Ala Leu Thr Ala Arg Asn Asn Gly Ala Gln Gly Ile Gly Leu Cys

620 625 630

CGG ACA GAG CAC ATG TTC TTT GCT TCA GAC GAG AGG ATT AAG GCT GTC 1973

Arg Thr Glu His Met Phe Phe Ala Ser Asp Glu Arg Ile Lys Ala Val

635 640 645 650

AGG CAG ATG ATT ATG GCT CCC ACG CTT GAG CTG AGG CAG CAG GCG CTC 2021

Arg Gln Met Ile Met Ala Pro Thr Leu Glu Leu Arg Gln Gln Ala Leu

655 660 665

GAC CGT CTC TTG ACG TAT CAG AGG TCT GAC TTC GAA GGC ATT TTC CGT 2069

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

670 675 680

GCT ATG GAT GGA CTC CCG GTG ACC ATC CGA CTC CTG GAC CAT CCT TCT 2117

Ala Met Asp Gly Leu Pro Val Thr Ile Arg Leu Leu Asp His Pro Ser

685 690 695

TAC GAG TTC CTT CCA GAA GGG AAC ATC GAG GAC ATT GTA AGT GAA TTA 2165

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

700 705 710

TGT GCT GAG ACG GGA GCC AAC CAG GAG GAT GCC CTC GCG CGA ATT GAA 2213

Cys Ala Glu Thr Gly Ala Asn Gln Glu Asp Ala Leu Ala Arg Ile Glu

715 720 725 730

AAG CTT TCA GAA GTA AAC CCG ATG CTT GGC TTC CGT GGG TGC AGG CTT 2261

Lys Leu Ser Glu Val Asn Pro Met Leu Gly Phe Arg Gly Cys Arg Leu

735 740 745

GGT ATA TCG TAC CCT GAA TTG ACA GAG ATG CAA GCC CGG GCC ATT TTT 2309

Gly Ile Ser Tyr Pro Glu Leu Thr Glu Met Gln Ala Arg Ala Ile Phe

750 755 760

GAA GCT GCT ATA GCA ATG ACC AAC CAG GGT GTT CAA GTG TTC CCA GAG 2357

Glu Ala Ala Ile Ala Met Thr Asn Gln Gly Val Gln Val Phe Pro Glu

765 770 775

ATA ATG GTT CCT CTT GTT GGA ACA CCA CAG GAA CTG GGG CAT CAA GTG 2405

Ile Met Val Pro Leu Val Gly Thr Pro Gln Glu Leu Gly His Gln Val

780 785 790

ACT CTT ATC CGC CAA GTT GCT GAG AAA GTG TTC GCC AAT GTG GGC AAG 2453

Thr Leu Ile Arg Gln Val Ala Glu Lys Val Phe Ala Asn Val Gly Lys

795 800 805 810

ACT ATC GGG TAC AAA GTT GGA ACA ATG ATT GAG ATC CCC AGG GCA GCT 2501

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

815 820 825

CTG GTG GCT GAT GAG ATA GCG GAG CAG GCT GAA TTC TTC TCC TTC GGA 2549

Leu Val Ala Asp Glu Ile Ala Glu Gln Ala Glu Phe Phe Ser Phe Gly

830 835 840

ACG AAC GAC CTG ACG CAG ATG ACC TTT GGG TAC AGC AGG GAT GAT GTG 2597

Thr Asn Asp Leu Thr Gln Met Thr Phe Gly Tyr Ser Arg Asp Asp Val

845 850 855

GGA AAG TTC ATT CCC GTT CAT CTT GCT CAG GGC ATC CTC CAA CAT GAC 2645

Gly Lys Phe Ile Pro Val His Leu Ala Gln Gly Ile Leu Gln His Asp

860 865 870

CCC TTC GAG GTC CTG GAC CAG AGG GGA GTG GGC GAG CTG GTG AAG TTT 2693

Pro Phe Glu Val Leu Asp Gln Arg Gly Val Gly Glu Leu Val Lys Phe

875 880 885 890

GCT ACA GAG AGG GGC CGC AAA GCT AGG CCT AAC TTG AAG GTG GGC ATT 2741

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

895 900 905

TGT GGA GAA CAC GGT GGA GAG CCT TCG TCT GTG GCC TTC TTC GCG AAG 2789

Cys Gly Glu His Gly Gly Glu Pro Ser Ser Val Ala Phe Phe Ala Lys

910 915 920

GCT GGG CTG GAT TTC GTT TCT TGC TCC CCT TTC AGG GTT CCG ATT GCT 2837

Ala Gly Leu Asp Phe Val Ser Cys Ser Pro Phe Arg Val Pro Ile Ala

925 930 935

AGG CTA GCT GCA GCT CAG GTG CTT GTC TGAGGCTGCC TCCTCG 2880

Arg Leu Ala Ala Ala Gln Val Leu Val

940 945

Sequence number: 3

Sequence is long: 2610

Sequence type: nucleic acid

Sequence

GAATTCTCAA TCCTTTGCTC ATCGCAGCAT ATCAATGTTA ACACATAAAC TTTAGGAGGA 60

AGAAAACTT ATG GCA AAA TGG GTT TAT AAG TTC GAA GAA GGC AAT GCA TCT 111

Met Ala Lys Trp Val Tyr Lys Phe Glu Glu Gly Asn Ala Ser

1 5 10

ATG AGA AAC CTT CTT GGA GGC AAA GGC TGC AAC CTT GCA GAG ATG ACC 159

Met Arg Asn Leu Leu Gly Gly Lys Gly Cys Asn Leu Ala Glu Met Thr

15 20 25 30

ATC TTA GGA ATG CCG ATT CCA CAG GGC TTT ACT GTA ACA ACA GAA GCT 207

Ile Leu Gly Met Pro Ile Pro Gln Gly Phe Thr Val Thr Thr Glu Ala

35 40 45

TGT ACA GAG TAC TAC AAC AGT GGA AAA CAG ATC ACA CAG GAA ATT CAG 255

Cys Thr Glu Tyr Tyr Asn Ser Gly Lys Gln Ile Thr Gln Glu Ile Gln

50 55 60

GAT CAG ATT TTC GAA GCT ATC ACA TGG TTA GAG GAA CTG AAC GGC AAG 303

Asp Gln Ile Phe Glu Ala Ile Thr Trp Leu Glu Glu Leu Asn Gly Lys

65 70 75

AAG TTC GGC GAC ACT GAA GAT CCG TTA TTA GTA TCT GTA CGT TCC GCG 351

Lys Phe Gly Asp Thr Glu Asp Pro Leu Leu Val Ser Val Arg Ser Ala

80 85 90

GCC CGC GCA TCC ATG CCG GGT ATG ATG GAT ACC ATC CTG AAC CTT GGT 399

Ala Arg Ala Ser Met Pro Gly Met Met Asp Thr Ile Leu Asn Leu Gly

95 100 105 110

TTA AAC GAC GTT GCA GTA GAG GGC TTT GCA AAG AAA ACG GGA AAT CCA 447

Leu Asn Asp Val Ala Val Glu Gly Phe Ala Lys Lys Thr Gly Asn Pro

115 120 125

AGA TTT GCA TAT GAT TCT TAC AGA AGA TTT ATC CAG ATG TAT TCC GAC 495

Arg Phe Ala Tyr Asp Ser Tyr Arg Arg Phe Ile Gln Met Tyr Ser Asp

130 135 140

GTA GTT ATG GAA GTT CCG AAG TCC CAT TTC GAG AAA ATC ATC GAT GCG 543

Val Val Met Glu Val Pro Lys Ser His Phe Glu Lys Ile Ile Asp Ala

145 150 155

ATG AAA GAA GAA AAG GGC GTT CAC TTC GAT ACA GAC CTG ACT GCC GAT 591

Met Lys Glu Glu Lys Gly Val His Phe Asp Thr Asp Leu Thr Ala Asp

160 165 170

GAT TTA AAA GAG CTG GCT GAG AAG TTC AAA GCT GTT TAC AAA GAG GCT 639

Asp Leu Lys Glu Leu Ala Glu Lys Phe Lys Ala Val Tyr Lys Glu Ala

175 180 185 190

ATG AAC GGC GAA GAG TTC CCA CAG GAG CCG AAG GAT CAG TTA ATG GGC 687

Met Asn Gly Glu Glu Phe Pro Gln Glu Pro Lys Asp Gln Leu Met Gly

195 200 205

GCT GTT AAA GCA GTT TTC CGT TCC TGG GAC AAC CCT CGT GCA ATC GTA 735

Ala Val Lys Ala Val Phe Arg Ser Trp Asp Asn Pro Arg Ala Ile Val

210 215 220

TAC CGC CGT ATG AAC GAT ATC CCT GGA GAC TGG GGT ACT GCA GTT AAC 783

Tyr Arg Arg Met Asn Asp Ile Pro Gly Asp Trp Gly Thr Ala Val Asn

225 230 235

GTT CAG ACC ATG GTA TTT GGT AAC AAG GGC GAG ACC AGC GGT ACA GGC 831

Val Gln Thr Met Val Phe Gly Asn Lys Gly Glu Thr Ser Gly Thr Gly

240 245 250

GTT GCC TTC ACA CGT AAC CCA TCC ACA GGT GAA AAA GGC ATC TAC GGT 879

Val Ala Phe Thr Arg Asn Pro Ser Thr Gly Glu Lys Gly Ile Tyr Gly

255 260 265 270

GAG TAC CTG ATC AAT GCA CAG GGC GAG GAC GTA GTT GCA GGT GTC CGC 927

Glu Tyr Leu Ile Asn Ala Gln Gly Glu Asp Val Val Ala Gly Val Arg

275 280 285

ACA CCA CAG CCT ATC ACC CAG TTA GAG AAC GAT ATG CCT GAC TGC TAC 975

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

290 295 300

AAG CAG TTC ATG GAT CTG GCC ATG AAG CTG GAG AAA CAT TTC CGT GAC 1023

Lys Gln Phe Met Asp Leu Ala Met Lys Leu Glu Lys His Phe Arg Asp

305 310 315

ATG CAG GAT ATG GAG TTC ACA ATC GAG GAA GGT AAA TTA TAC TTC TTA 1071

Met Gln Asp Met Glu Phe Thr Ile Glu Glu Gly Lys Leu Tyr Phe Leu

320 325 330

CAG ACA CGT AAC GGC AAG AGA ACA GCT CCG GCT GCT CTT CAG ATT GCC 1119

Gln Thr Arg Asn Gly Lys Arg Thr Ala Pro Ala Ala Leu Gln Ile Ala

335 340 345 350

TGC GAT TTA GTA GAC GAA GGC ATG ATC ACA GAG GAA GAG GCT GTT GTA 1167

Cys Asp Leu Val Asp Glu Gly Met Ile Thr Glu Glu Glu Ala Val Val

355 360 365

AGA ATC GAA GCA AAA TCT CTT GAT CAG TTA CTT CAC CCG ACC TTC AAC 1215

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

370 375 380

CCG GCT GCT TTA AAG GCC GGC GAA GTA ATC GGT TCC GCT CTT CCG GCA 1263

Pro Ala Ala Leu Lys Ala Gly Glu Val Ile Gly Ser Ala Leu Pro Ala

385 390 395

TCT CCT GGC GCA GCA GCA GGT AAA GTA TAC TTC ACC GCT GAT GAG GCT 1311

Ser Pro Gly Ala Ala Ala Gly Lys Val Tyr Phe Thr Ala Asp Glu Ala

400 405 410

AAG GCT GCC CAC GAG AAG GGT GAG AGA GTT ATC CTT GTT CGT CTT GAG 1359

Lys Ala Ala His Glu Lys Gly Glu Arg Val Ile Leu Val Arg Leu Glu

415 420 425 430

ACA TCT CCG GAA GAT ATC GAA GGT ATG CAT GCA GCC GAA GGT ATC CTG 1407

Thr Ser Pro Glu Asp Ile Glu Gly Met His Ala Ala Glu Gly Ile Leu

435 440 445

ACA GTG CGC GGC GGT ATG ACA AGC CAT GCA GCC GTA GTT GCA CGT GGT 1455

Thr Val Arg Gly Gly Met Thr Ser His Ala Ala Val Val Ala Arg Gly

450 455 460

ATG GGA ACA TGC TGC GTA TCC GGA TGC GGT GAG ATC AAG ATC AAC GAA 1503

Met Gly Thr Cys Cys Val Ser Gly Cys Gly Glu Ile Lys Ile Asn Glu

465 470 475

GAA GCT AAG ACA TTC GAA CTT GGC GGA CAC ACA TTT GCA GAG GGA GAT 1551

Glu Ala Lys Thr Phe Glu Leu Gly Gly His Thr Phe Ala Glu Gly Asp

480 485 490

TAC ATC TCC TTA GAT GGT TCC ACA GGT AAG ATT TAC AAG GGC GAC ATC 1599

Tyr Ile Ser Leu Asp Gly Ser Thr Gly Lys Ile Tyr Lys Gly Asp Ile

495 500 505 510

GAG ACT CAG GAA CGT TCC GTA AGC GGA AGC TTC GAG CGT ATC ATG GTA 1647

Glu Thr Gln Glu Arg Ser Val Ser Gly Ser Phe Glu Arg Ile Met Val

515 520 525

TGG GCT GAC AAG TTC AGA ACA TTA AAG GTT CGT ACA AAT GCC GAC ACA 1695

Trp Ala Asp Lys Phe Arg Thr Leu Lys Val Arg Thr Asn Ala Asp Thr

530 535 540

CCG GAA GAT ACA CTC AAT GCC GTT AAA CTG GGT GCA GAG GGC ATC GGT 1743

Pro Glu Asp Thr Leu Asn Ala Val Lys Leu Gly Ala Glu Gly Ile Gly

545 550 555

CTT TGC CGT ACA GAG CAT ATG TTC TTC GAG GCT GAC AGA ATC ATG AAG 1791

Leu Cys Arg Thr Glu His Met Phe Phe Glu Ala Asp Arg Ile Met Lys

560 565 570

ATC AGA AAG ATG ATC CTT TCC GAT TCA GTG GAA GCA AGA GAA GAG GCT 1839

Ile Arg Lys Met Ile Leu Ser Asp Ser Val Glu Ala Arg Glu Glu Ala

575 580 585 590

CTG AAC GAA TTA ATC CCG TTC CAG AAG GGC GAT TTC AAG GCT ATG TAC 1887

Leu Asn Glu Leu Ile Pro Phe Gln Lys Gly Asp Phe Lys Ala Met Tyr

595 600 605

AAA GCT CTG GAA GGC AGG CCA ATG ACG GTT CGC TAC CTG GAT CCG CCG 1935

Lys Ala Leu Glu Gly Arg Pro Met Thr Val Arg Tyr Leu Asp Pro Pro

610 615 620

CTG CAT GAG TTC GTT CCT CAT ACA GAA GAG GAG CAG GCT GAA CTG GCT 1983

Leu His Glu Phe Val Pro His Thr Glu Glu Glu Gln Ala Glu Leu Ala

625 630 635

AAG AAC ATG GGC CTT ACT TTA GCA GAA GTA AAA GCA AAA GTT GAC GAA 2031

Lys Asn Met Gly Leu Thr Leu Ala Glu Val Lys Ala Lys Val Asp Glu

640 645 650

TTA CAC GAG TTC AAC CCA ATG ATG GGC CAT CGT GGC TGC CGT CTT GCA 2079

Leu His Glu Phe Asn Pro Met Met Gly His Arg Gly Cys Arg Leu Ala

655 660 665 670

GTT ACC TAT CCG GAA ATT GCA AAG ATG CAG ACA AGA GCC GTT ATG GAA 2127

Val Thr Tyr Pro Glu Ile Ala Lys Met Gln Thr Arg Ala Val Met Glu

675 680 685

GCT GCT ATC GAA GTG AAG GAA GAG ACA GGA ATC GAT ATT GTT CCT GAG 2175

Ala Ala Ile Glu Val Lys Glu Glu Thr Gly Ile Asp Ile Val Pro Glu

690 695 700

ATC ATG ATT CCG TTA GTT GGC GAG AAG AAA GAG CTT AAG TTC GTT AAG 2223

Ile Met Ile Pro Leu Val Gly Glu Lys Lys Glu Leu Lys Phe Val Lys

705 710 715

GAC GTA GTT GTG GAA GTA GCT GAG CAG GTT AAG AAA GAG AAA GGT TCC 2271

Asp Val Val Val Glu Val Ala Glu Gln Val Lys Lys Glu Lys Gly Ser

720 725 730

GAT ATG CAG TAC CAC ATC GGT ACC ATG ATC GAA ATT CCT CGT GCA GCT 2319

Asp Met Gln Tyr His Ile Gly Thr Met Ile Glu Ile Pro Arg Ala Ala

735 740 745 750

CTC ACA GCA GAT GCC ATC GCT GAG GAA GCA GAG TTC TTC TCC TTC GGT 2367

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

755 760 765

ACA AAC GAC TTA ACA CAG ATG ACA TTC GGC TTC TCC CGT GAC GAC GCC 2415

Thr Asn Asp Leu Thr Gln Met Thr Phe Gly Phe Ser Arg Asp Asp Ala

770 775 780

GGC AAG TTC CTG GAT TCC TAC TAT AAA GCA AAA ATT TAT GAG TCC GAT 2463

Gly Lys Phe Leu Asp Ser Tyr Tyr Lys Ala Lys Ile Tyr Glu Ser Asp

785 790 795

CCA TTC GCA AGA CTT GAC CAG ACA GGC GTT GGC CAG TTA GTA GAG ATG 2511

Pro Phe Ala Arg Leu Asp Gln Thr Gly Val Gly Gln Leu Val Glu Met

800 805 810

GCA GTT AAG AAA GGC CGT CAG ACA CGT CCG GGC CTT AAG TGC GGC ATC 2559

Ala Val Lys Lys Gly Arg Gln Thr Arg Pro Gly Leu Lys Cys Gly Ile

815 820 825 830

TGC GGC GAG CAC GGC GAG ATC CTT CTT CCG TAGAGTTCTG CCACAAAGTA 2609

Cys Gly Glu His Gly Glu Ile Leu Leu Pro

835 840

G 2610

Sequence number: 4

Sequence is long: 2722

Sequence type: nucleic acid

Sequence

GAACTATTTA AGGAATTTGT AAGAATTTAG AGTTCATTCA GATAATA ATG CAA AGA 56

Met Gln Arg

1

GTA TAT GCT TTC GAA GAT GGT GAT GGA ACC AAC AAG AAA CTC CTT GGA 104

Val Tyr Ala Phe Glu Asp Gly Asp Gly Thr Asn Lys Lys Leu Leu Gly

5 10 15

GGA AAG GGA GCT GGA CTT TGC ACA ATG ACA AAA ATT GGA CTT CCA GTT 152

Gly Lys Gly Ala Gly Leu Cys Thr Met Thr Lys Ile Gly Leu Pro Val

20 25 30 35

CCA CAA GGA TTT GTT ATT ACA ACT GAA ATG TGT AAA CAA TTC ATT GCT 200

Pro Gln Gly Phe Val Ile Thr Thr Glu Met Cys Lys Gln Phe Ile Ala

40 45 50

AAT GGA AAC AAA ATG CCA GAA GGA TTA ATG GAA GAA GTT AAA AAA GAA 248

Asn Gly Asn Lys Met Pro Glu Gly Leu Met Glu Glu Val Lys Lys Glu

55 60 65

TAT CAA TTA GTT GAA AAG AAA TCA GGA AAA GTC TTT GGA GGA GAA GAA 296

Tyr Gln Leu Val Glu Lys Lys Ser Gly Lys Val Phe Gly Gly Glu Glu

70 75 80

AAT CCA CTT CTT GTT TCA GTC AGA TCA GGA GCT GCT ATG TCT ATG CCA 344

Asn Pro Leu Leu Val Ser Val Arg Ser Gly Ala Ala Met Ser Met Pro

85 90 95

GGT ATG ATG GAT ACT ATT CTT AAT CTT GGA CTT AAT GAT AAA ACT GTT 392

Gly Met Met Asp Thr Ile Leu Asn Leu Gly Leu Asn Asp Lys Thr Val

100 105 110 115

GTT GCT CTT GCT AAA TTA ACC AAC AAT GAA AGA TTT GCA TAT GAT TCA 440

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

120 125 130

TAC AGA AGA TTT GTT TCC CTC TTC GGA AAG ATT GCT CTT AAT GCT TGT 488

Tyr Arg Arg Phe Val Ser Leu Phe Gly Lys Ile AlaLeu Asn Ala Cys

135 140 145

GAT GAA GTT TAT GAT AAG ACT CTT GAA AAC AAA AAA GTT GAA AAG GGA 536

Asp Glu Val Tyr Asp Lys Thr Leu Glu Asn Lys Lys Val Glu Lys Gly

150 155 160

GTT AAA TTA GAT ACT GAA TTA GAT GCT AAT GAT ATG AAA GAA CTT GCA 584

Val Lys Leu Asp Thr Glu Leu Asp Ala Asn Asp Met Lys Glu Leu Ala

165 170 175

CAA GTC TTC ATT AAA AAG ACT GAA GAA TTC ACT AAA CAA CCA TTC CCA 632

Gln Val Phe Ile Lys Lys Thr Glu Glu Phe Thr Lys Gln Pro Phe Pro

180 185 190 195

GTT GAT CCA TAT GCT CAA TTA GAA TTT GCC ATT TGT GCT GTA TTC AGA 680

Val Asp Pro Tyr Ala Gln Leu Glu Phe Ala Ile Cys Ala Val Phe Arg

200 205 210

TCA TGG ATG GGA AAG AGA GCT GTT GAT TAC AGA AGA GAA TTC AAG ATT 728

Ser Trp Met Gly Lys Arg Ala Val Asp Tyr Arg Arg Glu Phe Lys Ile

215 220 225

ACT CCA GAA CAA GCT GAT GGA ACT GCT GTT TCA GTT GTT TCT ATG GTT 776

Thr Pro Glu Gln Ala Asp Gly Thr Ala Val Ser Val Val Ser Met Val

230 235 240

TAT GGT AAT ATG GGT AAT GAT TCA GCT ACT GGT GTT TGT TTC ACT AGA 824

Tyr Gly Asn Met Gly Asn Asp Ser Ala Thr Gly Val Cys Phe Thr Arg

245 250 255

GAT CCA GGA ACA GGA GAA AAT ATG TTC TTC GGA GAA TAT CTT AAG AAT 872

Asp Pro Gly Thr Gly Glu Asn Met Phe Phe Gly Glu Tyr Leu Lys Asn

260 265 270 275

GGA CAA GGA GAA GAT GTT GTT GCT GGT ATT AGA ACA CCA CAA ATT ATT 920

Ala Gln Gly Glu Asp Val Val Ala Gly Ile Arg Thr Pro Gln Ile Ile

280 285 290

TCA AAG ATG GCA GAA GAT CGA GAT CTT CCA GGT TGC TAT GAA CAA CTT 968

Ser Lys Met Ala Glu Asp Arg Asp Leu Pro Gly Cys Tyr Glu Gln Leu

295 300 305

CTT GAT ATT AGA AAG AAA TTA GAA GGA TAT TTC CAT GAA GTA CAA GAC 1016

Leu Asp Ile Arg Lys Lys Leu Glu Gly Tyr Phe His Glu Val Gln Asp

310 315 320

TTT GAA TTC ACT ATT GAA AGA AAG AAA CTT TAC ATG CTC CAA ACT AGA 1064

Phe Glu Phe Thr Ile Glu Arg Lys Lys Leu Tyr Met Leu Gln Thr Arg

325 330 335

AAT GGA AAG ATG AAT GCA ACT GCT ACT GTC AGA ACA GGA GTT GAT ATG 1112

Asn Gly Lys Met Asn Ala Thr Ala Thr Val Arg Thr Gly Val Asp Met

340 345 350 355

GTT GAA GAA GGA CTT ATT ACA AAA GAA CAA GCC ATT ATG AGA ATT GCA 1160

Val Glu Glu Gly Leu Ile Thr Lys Glu Gln Ala Ile Met Arg Ile Ala

360 365 370

CCA CAA TCA GTT GAT CAA TTA CTT CAT AAG AAT ATG CCA GCT AAT TAT 1208

Pro Gln Ser Val Asp Gln Leu Leu His Lys Asn Met Pro Ala Asn Tyr

375 380 385

GCA GAA GCT CCA TTA GTT AAA GGA CTT CCA GCA TCA CCA GGA GCT GCT 1256

Ala Glu Ala Pro Leu Val Lys Gly Leu Pro Ala Ser Pro Gly Ala Ala

390 395 400

ACA GGA GCT GTT GTT TTT GAT GCC GAT GAT GCA GTT GAA CAA GCT AAA 1304

Thr Gly Ala Val Val Phe Asp Ala Asp Asp Ala Val Glu Gln Ala Lys

405 410 415

GGA AAG AAA GTT CTT CTT CTT AGA GAA GAA ACT AAA CCA GAA GAT ATT 1352

Gly Lys Lys Val Leu Leu Leu Arg Glu Glu Thr Lys Pro Glu Asp Ile

420 425 430 435

CAT GGA TTC TTT GTT GCT GAA GGT ATT TTA ACC TGC AGA GGA GGA AAA 1400

His Gly Phe Phe Val Ala Glu Gly Ile Leu Thr Cys Arg Gly Gly Lys

440 445 450

ACA TCA CAC GCA GCT GTC GTT GCT AGA GGT ATG GGT AAA CCA TGT GTT 1448

Thr Ser His Ala Ala Val Val Ala Arg Gly Met Gly Lys Pro Cys Val

455 460 465

TCA GGA GCT GAA GGA ATT AAA GTT GAT GTT GCT AAG AAA ATT GCT AAG 1496

Ser Gly Ala Glu Gly Ile Lys Val Asp Val Ala Lys Lys Ile Ala Lys

470 475 480

ATT GGA AGC CTT GAA GTT CAT GAA GGA GAT ATT TTA ACT ATT GAT GGA 1544

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

485 490 495

TCA ACT GGA TGT GTC TAT AAG GGA GAA GTT CCA TTA GAA GAA CCA CAA 1592

Ser Thr Gly Cys Val Tyr Lys Gly Glu Val Pro Leu Glu Glu Pro Gln

500 505 510 515

GTT GGA TCA GGA TAT TTC GGA ACC ATC TTA AAA TGG GCC AAT GAA ATT 1640

Val Gly Ser Gly Tyr Phe Gly Thr Ile Leu Lys Trp Ala Asn Glu Ile

520 525 530

AAA AAG ATT GGA GTT TTT GCT GCT GGA GAT CTT CCA TCA GCT GCT AAG 1688

Lys Lys Ile Gly Val Phe Ala Ala Gly Asp Leu Pro Ser Ala Ala Lys

535 540 545

AAA GCC CTT GAA TTT GGA GCT GAA GGT ATT GGA CTT TGC AGA ACT GAA 1736

Lys Ala Leu Glu Phe Gly Ala Glu Gly Ile Gly Leu Cys Arg Thr Glu

550 555 560

CGT ATG TTC AAT GCA GTT GAA AGA CTT CCA ATT GTT GTC AAG ATG ATT 1784

Arg Met Phe Asn Ala Val Glu Arg Leu Pro Ile Val Val Lys Met Ile

565 570 575

CTT TCA AAT ACC CTT GAA GAA AGA AAG AAA TAT CTT AAT GAA CTT ATG 1832

Leu Ser Asn Thr Leu Glu Glu Arg Lys Lys Tyr Leu Asn Glu Leu Met

580 585 590 595

CCA CTT CAA AAA CAA GAT TTC ATT GGA TTA TTG AAG ACT ATG AAT GGA 1880

Pro Leu Gln Lys Gln Asp Phe Ile Gly Leu Leu Lys Thr Met Asn Gly

600 605 610

CTT CCA GTC ACT GTC AGA CTT CTT GAT CCA CCA TTA CAT GAA TTC CTC 1928

Leu Pro Val Thr Val Arg Leu Leu Asp Pro Pro Leu His Glu Phe Leu

615 620 625

CCA ACT CTT GAA GAG TTA ATG AGA GAA ATC TTT GAA ATG AAA CTT TCA 1976

Pro Thr Leu Glu Glu Leu Met Arg Glu Ile Phe Glu Met Lys Leu Ser

630 635 640

GGT AAG AGT GAA GGA CTT GCA GAA AAA GAA GTT GTT CTT AAG AAA GTT 2024

Gly Lys Thr Glu Gly Leu Ala Glu Lys Glu Val Val Leu Lys Lys Val

645 650 655

AAA GAA CTT ATG GAA GTT AAT CCA ATG ATT GGA CAC AGA GGA ATT AGA 2072

Lys Glu Leu Met Glu Val Asn Pro Met Ile Gly His Arg Gly Ile Arg

660 665 670 675

CTT GGA ACT ACT AAT CCA GAA ATT TAT GAA ATG CAA ATT AGA GCA TTC 2120

Leu Gly Thr Thr Asn Pro Glu Ile Tyr Glu Met Gln Ile Arg Ala Phe

680 685 690

TTA GAA GCT ACT CGT GAA GTT ATT AAG GAA GGA ATT AAC GAT CAT CGA 2168

Leu Glu Ala Thr Arg Glu Val Ile Lys Glu Gly Ile Asn Asp His Arg

695 700 705

GAA ATT ATG ATT CCA AAT GTT ACA GAA GTT AAT GAA CTT ATT AAC TTA 2216

Glu Ile Met Ile Pro Asn Val Thr Glu Val Asn Glu Leu Ile Asn Leu

710 715 720

AGA AAG AAT GTT CTT GAA CCA GTT CAT GAA GAA GTT GAA AAG AAA TAT 2264

Arg Lys Asn Val Leu Glu Pro Val His Glu Glu Val Glu Lys Lys Tyr

725 730 735

GGT ATT AAA GTA CCA TTC TCG TAT GGT ACT ATG GTT GAA TGT GTT AGA 2312

Gly Ile Lys Val Pro Phe Ser Tyr Gly Thr Met Val Glu Cys Val Arg

740 745 750 755

GCA GCA TTA ACA GCT GAT AAG ATT GCT ACA GAA GCT TCA TTC TTC TCA 2360

Ala Ala Leu Thr Ala Asp Lys Ile Ala Thr Glu Ala Ser Phe Phe Ser

760 765 770

TTC GGA ACT AAT GAT CTT ACA CAA GGA ACA TTC TCA TAC TCA CGT GAA 2408

Phe Gly Thr Asn Asp Leu Thr Gln Gly Thr Phe Ser Tyr Ser Arg Glu

775 780 785

GAT TCA GAA AAC AAA TTC ATT CCA AAA TAT GTT GAA CTT AAG ATT CTT 2456

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

790 795 800

CCA GCT AAT CCA TTT GAA ATT CTT GAT AGA CCA GGT GTT GGA GAA GTT 2504

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

805 810 815

ATG AGA ATT GCT GTT ACT AAA GGA AGA CAA ACA AGA CCA GAA TTA CTT 2552

Met Arg Ile Ala Val Thr Lys Gly Arg Gln Thr Arg Pro Glu Leu Leu

820 825 830 835

GTT GGT ATT TGT GGA GAA CAC GGA GGA GAA CCA TCA TCA ATT GAA TGG 2600

Val Gly Ile Cys Gly Glu His Gly Gly Glu Pro Ser Ser Ile Glu Trp

840 845 850

TGC CAC ATG ATT GGA TTG AAC TAT GTT TCA TGT TCT TCA TAC AGA ATT 2648

Cys His Met Ile Gly Leu Asn Tyr Val Ser Cys Ser Ser Tyr Arg Ile

855 860 865

CCA GTT GCT AGA ATT GCT GCT GCT CAA GCC CAA ATT AGA CAT CCA AGA 2696

Pro Val Ala Arg Ile Ala Ala Ala Gln Ala Gln Ile Arg His Pro Arg

870 875 880

GAA AAT TAAATTAACT TTTTTGGTTT 2722

Glu Asn

885

Sequence number: 5

Sequence is long: 3180

Sequence type: nucleic acid

Sequence

CTGAAATTCC CGTAATCTAT CATCATTTAC ACCACAAATC GATTCACATC CTCACCGAAT 60

AGAAATCAAA TATCATTTAC TCCATCTCAC GATCTCCTTT TGCTATTGCT GATACCTCAA 120

TTTCGCAGGT GAAGGCGGAC G ATG AGT TCG TTG TTT GTT GAA GGT ATG CCT 171

Met Ser Ser Leu Phe Val Glu Gly Met Pro

1 5 10

CTG AAG TCA GCC AAT GAG TCG TGC TTA CCG GCG AGC GTG AAG CAA CGG 219

Leu Lys Ser Ala Asn Glu Ser Cys Leu Pro Ala Ser Val Lys Gln Arg

15 20 25

CGA ACC GGT GAT CTC AGG CGA TTG AAC CAC CAC CGT CAA CCG GCG TTT 267

Arg Thr Gly Asp Leu Arg Arg Leu Asn His His Arg Gln Pro Ala Phe

30 35 40

GTC CGG GGG ATT TGC CGT CGG AAG TTG AGT GGA GTT AGC AGA ATA GAG 315

Val Arg Gly Ile Cys Arg Arg Lys Leu Ser Gly Val Ser Arg Ile Glu

45 50 55

TTG CGC ACC GGT GGT TTA ACT CTG CCA CGA GCG GTG CTT AAT CCG GTG 363

Leu Arg Thr Gly Gly Leu Thr Leu Pro Arg Ala Val Leu Asn Pro Val

60 65 70

TCT CCT CCG GTA ACG ACG ACT AAA AAG AGG GTT TTC ACT TTT GGT AAA 411

Ser Pro Pro Val Thr Thr Thr Lys Lys Arg Val Phe Thr Phe Gly Lys

75 80 85 90

GGA AAC AGT GAA GGC AAC AAG GAC ATG AAA TCC TTG TTG GGA GGA AAA 459

Gly Asn Ser Glu Gly Asn Lys Asp Met Lys Ser Leu Leu Gly Gly Lys

95 100 105

GGT GCA AAT CTT GCA GAG ATG GCA AGC ATT GGC CTA TCA GTT CCT CCT 507

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

110 115 120

GGG CTC ACT ATT TCA ACT GAA GCA TGT GAG GAA TAT CAA CAA AAT GGA 555

Gly Leu Thr Ile Ser Thr Glu Ala Cys Glu Glu Tyr Gln Gln Asn Gly

125 130 135

AAA AAA CTG CCT CCA GGT TTA TGG GAT GAG ATT CTG GAA GGC TTA CAG 603

Lys Lys Leu Pro Pro Gly Leu Trp Asp Glu Ile Leu Glu Gly Leu Gln

140 145 150

TAT GTC CAG AAA GAG ATG TCT GCA TCT CTC GGT GAC CCG TCT AAA GCT 651

Tyr Val Gln Lys Glu Met Ser Ala Ser Leu Gly Asp Pro Ser Lys Ala

155 160 165 170

CTC CTC CTT TCC GTC CGT TCG GGT GCT GCC ATA TCG ATG CCT GGT ATG 699

Leu Leu Leu Ser Val Arg Ser Gly Ala Ala Ile Ser Met Pro Gly Met

175 180 185

ATG GAC ACT GTA TTG AAT CTC GGG CTT AAT GAT GAG GTC GTA GAT GGT 747

Met Asp Thr Val Leu Asn Leu Gly Leu Asn Asp Glu Val Val Asp Gly

190 195 200

CTA GCT GCC AAA AGT GGA GCT CGC TTT GCC TAT GAC TCG TAT AGG AGG 795

Leu Ala Ala Lys Ser Gly Ala Arg Phe Ala Tyr Asp Ser Tyr Arg Arg

205 210 215

TTT CTA GAT ATG TTT GGC AAC GTT GTA ATG GGT ATC CCA CAT TCG TTA 843

Phe Leu Asp Met Phe Gly Asn Val Val Met Gly Ile Pro His Ser Leu

220 225 230

TTT GAT GAA AAG TTA GAG CAG ATG AAA GCT GAA AAA GGG ATT CAT CTC 891

Phe Asp Glu Lys Leu Glu Gln Met Lys Ala Glu Lys Gly Ile His Leu

235 240 245 250

GAC ACT GAT CTC ACT GCT GCT GAT CTT AAA GAT CTT GCT GAG CAA TAC 939

Asp Thr Asp Leu Thr Ala Ala Asp Leu Lys Asp Leu Ala Glu Gln Tyr

255 260 265

AAG AAC GTG TAT GTG GAA GCA AAG GGC GAA AAG TTT CCC ACA GAT CCA 987

Lys Asn Val Tyr Val Glu Ala Lys Gly Glu Lys Phe Pro Thr Asp Pro

270 275 280

AAG AAA CAG CTA GAG TTA GCA GTG AAT GCG GTT TTT GAT TCT TGG GAC 1035

Lys Lys Gln Leu Glu Leu Ala Val Asn Ala Val Phe Asp Ser Trp Asp

285 290 295

AGC CCA AGG GCC AAT AAG TAC AGG AGT ATT AAC CAG ATA ACT GGG TTA 1083

Ser Pro Arg Ala Asn Lys Tyr Arg Ser Ile Asn Gln Ile Thr Gly Leu

300 305 310

AAG GGG ACC GCG GTT AAC ATT CAA TGC ATG GTG TTT GGC AAC ATG GGG 1131

Lys Gly Thr Ala Val Asn Ile Gln Cys Met Val Phe Gly Asn Met Gly

315 320 325 330

AAC ACT TCA GGA ACC GGT GTT CTT TTC ACT AGG AAC CCA AGC ACT GGT 1179

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

335 340 345

GAG AAG AAG CTG TAT GGG GAG TTT TTA GTC AAT GCT CAG GGA GAG GAT 1227

Glu Lys Lys Leu Tyr Gly Glu Phe Leu Val Asn Ala Gln Gly Glu Asp

350 355 360

GTT GTT GCT GGG ATC AGA ACA CCA GAA GAT TTG GTG ACC ATG GAG ACT 1275

Val Val Ala Gly Ile Arg Thr Pro Glu Asp Leu Val Thr Met Glu Thr

365 370 375

TGC ATG CCT GAA GCA TAC AGA GAG CTT GTG GAG AAC TGT GTG ATT TTA 1323

Cys Met Pro Glu Ala Tyr Arg Glu Leu Val Glu Asn Cys Val Ile Leu

380 385 390

GAG AGA CAC TAC AAA GAT ATG ATG GAT ATT GAA TTC ACA GTT CAA GAA 1371

Glu Arg His Tyr Lys Asp Met Met Asp Ile Glu Phe Thr Val Gln Glu

395 400 405 410

AAC AGA CTT TGG ATG CTG CAA TGC CGA ACA GGG AAA CGT ACT GGG AAA 1419

Asn Arg Leu Trp Met Leu Gln Cys Arg Thr Gly Lys Arg Thr Gly Lys

415 420 425

GGT GCG GTG AGA ATT GCA GTA GAT ATG GTG AAC GAA GGG CTA ATT GAT 1467

Gly Ala Val Arg Ile Ala Val Asp Met Val Asn Glu Gly Leu Ile Asp

430 435 440

ACT AGA ACA GCA ATT AAG AGG GTT GAG ACT CAA CAT CTA GAT CAG CTT 1515

Thr Arg Thr Ala Ile Lys Arg Val Glu Thr Gln His Leu Asp Gln Leu

445 450 455

CTT CAT CCA CAG TTT GAG AAT CCG TCT GCT TAC AAA AGC CAT GTG GTA 1563

Leu His Pro Gln Phe Glu Asn Pro Ser Ala Tyr Lys Ser His Val Val

460 465 470

GCA ACC GGT TTG CCA GCA TCC CCT GGG GCA GCC GTG GGG CAG GTT GTG 1611

Ala Thr Gly Leu Pro Ala Ser Pro Gly Ala Ala Val Gly Gln Val Val

475 480 485 490

TTC AGC GCA GAG GAT GCT GAA ACA TGG CAT GCA CAA GGA AAG AGT GCT 1659

Phe Ser Ala Glu Asp Ala Glu Thr Trp His Ala Gln Gly Lys Ser Ala

495 500 505

ATC TTG GTA AGG ACT GAA ACA AGC CCA GAA GAT GTT GGT GGT ATG CAT 1707

Ile Leu Val Arg Thr GluT hr Ser Pro Glu Asp Val Gly Gly Met His

510 515 520

GCA GCA GCT GGA ATC TTA ACC GCT AGA GGA GGA ATG ACA TCA CAT GCA 1755

Ala Ala Ala Gly Ile Leu Thr Ala Arg Gly Gly Met Thr Ser His Ala

525 530 535

GCA GTG GTG GCT CGC GGA TGG GGC AAA TGT TGT GTT TCT GGT TGT GCT 1803

Ala Val Val Ala Arg Gly Trp Gly Lys Cys Cys Val Ser Gly Cys Ala

540 545 550

GAT ATT CGT GTG AAC GAT GAT ATG AAG GTT TTT ACG ATA GGT GAC CGT 1851

Asp Ile Arg Val Asn Asp Asp Met Lys Val Phe Thr Ile Gly Asp Arg

555 560 565 570

GTG ATT AAA GAA GGT GAC TGG CTT TCA CTT AAT GGT TCA ACT GGC GAA 1899

Val Ile Lys Glu Gly Asp Trp Leu Ser Leu Asn Gly Ser Thr Gly Glu

575 580 585

GTC ATA TTG GGT AAA CAG CTA CTG GCT CCA CCT GCA ATG AGC AAT GAT 1947

Val Ile Leu Gly Lys Gln Leu Leu Ala Pro Pro Ala Met Ser Asn Asp

590 595 600

TTA GAA ACA TTC ATG TCA TGG GCT GAT CAA GCA AGG CGT CTC AAG GTT 1995

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

605 610 615

ATG GCA AAT GCA GAC ACA CCT AAT GAT GCA TTA ACA GCC AGA AAC AAT 2043

Met Ala Asn Ala Asp Thr Pro Asn Asp Ala Leu Thr Ala Arg Asn Asn

620 625 630

GGT GCA CAA GGG ATC GGA CTC TGT AGA ACT GAA CAT ATG TTT TTC GCT 2091

Gly Ala Gln Gly Ile Gly Leu Cys Arg Thr Glu His Met Phe Phe Ala

635 640 645 650

TCT GAC GAG AGG ATC AAA GCT GTA AGA AAG ATG ATC ATG GCG GTC ACT 2139

Ser Asp Glu Arg Ile Lys Ala Val Arg Lys Met Ile Met Ala Val Thr

655 660 665

CCA GAA CAA AGA AAA GCG GCT CTA GAC CTC TTA CTC CCA TAC CAA AGA 2187

Pro Glu Gln Arg Lys Ala Ala Leu Asp Leu Leu Leu Pro Tyr Gln Arg

670 675 680

TCC GAT TTT GAG GGC ATT TTC CGA GCA ATG GAT GGA CTT CCT GTA ACA 2235

Ser Asp Phe Glu Gly Ile Phe Arg Ala Met Asp Gly Leu Pro Val Thr

685 690 695

ATC CGC CTT CTA GAC CCT CCA CTT CAT GAG TTT CTA CCC GAA GGT GAT 2283

Ile Arg Leu Leu Asp Pro Pro Leu His Glu Phe Leu Pro Glu Gly Asp

700 705 710

CTA GAA CAC ATA GTG AAT GAA CTT ACA GCG GAT ACA GGC ATG AGC AAA 2331

Leu Glu His Ile Val Asn Glu Leu Thr Ala Asp Thr Gly Met Ser Lys

715 720 725 730

GAT GAA ATC TAT TCA AGA ATC GAA AAA TTA TCC GAA GTG AAC CCT ATG 2379

Asp Glu Ile Tyr Ser Arg Ile Glu Lys Leu Ser Glu Val Asn Pro Met

735 740 745

CTT GGT TTC CGT GGT TGC CGA TTA GGG ATT TCA TAC CCC GAG CTA ACA 2427

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

750 755 760

GAA ATG CAA GTT CGT GCG ATC TTT CAA GCT GCA GTG TCT ATG AAC AAT 2475

Glu Met Gln Val Arg Ala Ile Phe Gln Ala Ala Val Ser Met Asn Asn

765 770 775

CAG GGG GTG ACT GTA ATA CCA GAG ATC ATG GTT CCG TTA GTC GGA ACA 2523

Gln Gly Val Thr Val Ile Pro Glu Ile Met Val Pro Leu Val Gly Thr

780 785 790

CCT CAG GAA TTA CGG CAT CAA ATC GGC GTA ATT CGT GGT GTA GCT GCA 2571

Pro Gln Glu Leu Arg His Gln Ile Gly Val Ile Arg Gly Val Ala Ala

795 800 805 810

AAT GTT TTT GCT GAA ATG GGG CTG ACG TTG GAG TAT AAA GTG GGA ACG 2619

Asn Val Phe Ala Glu Met Gly Leu Thr Leu Glu Tyr Lys Val Gly Thr

815 820 825

ATG ATT GAG ATT CCT CGA GCT GCT TTG ATT GCT GAT GAG ATT GCA AAA 2667

Met Ile Glu Ile Pro Arg Ala Ala Leu Ile Ala Asp Glu Ile Ala Lys

830 835 840

GAA GCC GAG TTC TTT TCG TTT GGA ACC AAT GAT TTG ACC CAG ATG ACA 2715

Glu Ala Glu Phe Phe Ser Phe Gly Thr Asn Asp Leu Thr Gln Met Thr

845 850 855

TTT GGG TAC AGC AGA GAT GAT GTT GGC AAG TTT TTG CCG ATT TAT CTT 2763

Phe Gly Tyr Ser Arg Asp Asp Val Gly Lys Phe Leu Pro Ile Tyr Leu

860 865 870

TCT CAA GGC ATT CTG CAG CAT GAT CCA TTT GAG GTT CTT GAC CAG AAA 2811

Ser Gln Gly Ile Leu Gln His Asp Pro Phe Glu Val Leu Asp Gln Lys

875 880 885 890

GGG GTG GGT CAA TTG ATC AAG ATG GCC ACG GAG AAA GGT CGT GCA GCC 2859

Gly Val Gly Gln Leu Ile Lys Met Ala Thr Glu Lys Gly Arg Ala Ala

895 900 905

AAT CCT AAC TTA AAG GTT GGG ATA TGT GGG GAG CAT GGT GGA GAA CCT 2907

Asn Pro Asn Leu Lys Val Gly Ile Cys Gly Glu His Gly Gly Glu Pro

910 915 920

TCT TCT GTT GCA TTT TTT GAC GGA GTT GGA CTA GAT TAT GTG TCG TGC 2955

Ser Ser Val Ala Phe Phe Asp Gly Val Gly Leu Asp Tyr Val Ser Cys

925 930 935

TCT CCA TTC AGG GTT CCT ATC GCA AGG TTG GCC GCT GCA CAA GTC GTT 3003

Ser Pro Phe Arg Val Pro Ile Ala Arg Leu Ala Ala Ala Gln Val Val

940 945 950

GTT TAAGCTTTGA AAGGAGGATG GCTTATTTGC TTCATGTTTT CCGCCATTGT 3056

Val

955

ATATTATTTT GGTTTCATCC TTATTGTAAT GGTGAAAATG AACGATGTTT AAACAAAACA 3116

ACCCATTATA TTTTGGTTTG GTATGCAATA ATCTACTTTT CAAACAAAAA AAAAAAAAAA 3176

AAAA 3180

Sequence number: 6

Sequence is long: 18

Sequence type: nucleic acid

Sequence

GACGGCTAAA AAGAGGGT 18

Sequence number: 7

Sequence is long: 20

Sequence type: nucleic acid

Sequence

TATCGAGAAA CCTTCTATAC 20

Sequence number: 8

Sequence is long: 17

Sequence type: nucleic acid

Sequence

GTTTTCCCAG TCACGAC 17

Sequence number: 9

Sequence is long: 17

Sequence type: nucleic acid

Sequence

CAGGAAACAG CTATGAC 17

Sequence number: 10

Sequence is long: 23

Sequence type: nucleic acid

Sequence

GATATCAATC CGGTGTCTCC TCC 23

Sequence number: 11

Sequence is long: 34

Sequence type: nucleic acid

Sequence

CGGTGTCTCC TCCGGATATC ACGGCTAAAA AGAG 34

Sequence number: 12

Sequence is long: 27

Sequence type: nucleic acid

Sequence

TTGATATCCC GGTTGTCTCC TCCGGTA 27

Sequence number: 13

Sequence is long: 20

Sequence type: nucleic acid

Sequence

GCAGAGATGA TGTTGGCAAG 20

Sequence number: 14

Sequence is long: 20

Sequence type: nucleic acid

Sequence

CTTGCCAACA TCATCTCTGC 20

Sequence number: 15

Sequence is long: 20

Sequence type: nucleic acid

Sequence

CTCACTGTTC GAAGAGAAGC 20

Sequence number: 16

Sequence is long: 23

Sequence type: nucleic acid

Sequence

CATATGCTCT GTCCGGCATA ATC 23

Sequence number: 17

Sequence is long: 21

Sequence type: nucleic acid

Sequence

CTCGAGGGAT CTCAATCATT G 21

Sequence number: 18

Sequence is long: 18

Sequence type: nucleic acid

Sequence

GCAATCTCTT CAGCAATC 18

Sequence number: 19

Sequence is long: 20

Sequence type: nucleic acid

Sequence

GCTTCTTTTC CAATCTCATC 20

Sequence number: 20

Sequence is long: 18

Sequence type: nucleic acid

Sequence

CGAAAAGAAA TCGGCTTC 18

Sequence number: 21

Sequence is long: 24

Sequence type: nucleic acid

Sequence

GAAAGATAAA TCTGCAAAAA CTTG 24

Sequence number: 22

Sequence is long: 19

Sequence type: nucleic acid

Sequence

GCCTTGAGCA AGATAAATC 19

Sequence number: 23

Sequence is long: 21

Sequence type: nucleic acid

Sequence

TTCTGGTCAA TAACCTCAAT G 21

Sequence number: 24

Sequence is long: 20

Sequence type: nucleic acid

Sequence

GCTTAAACAA TGACTTGTGC 20

Sequence number: 25

Sequence is long: 22

Sequence type: nucleic acid

Sequence

CCAATCTCAT CAGCTATTAA AG 22

Sequence number: 26

Sequence is long: 20

Sequence type: nucleic acid

Sequence

GCTTCTTTTG CAATCTCTTC 20

Sequence number: 27

Sequence is long: 18

Sequence type: nucleic acid

Sequence

CGAAAAGAAC TCAGCTTC 18

Sequence number: 28

Sequence is long: 23

Sequence type: nucleic acid

Sequence

CAAGATAAAT CGGCAAAAAC TTG 23

Sequence number: 29

Sequence is long: 23

Sequence type: nucleic acid

Sequence

GAATGCCTTG AGAAAGATAA ATC 23

Sequence number: 30

Sequence is long: 24

Sequence type: nucleic acid

Sequence

CTTTCTGGTC AAGAACCTCA AATG 24

Sequence number: 31

Sequence is long: 20

Sequence type: nucleic acid

Sequence

GCTTAAACAA CGACTTGTGC 20

Claims (7)

1. a peptide species, its aminoacid sequence is shown in sequence table sequence number 5.

2. a peptide species wherein contains the 832nd～No. 955 part of aminoacid sequence shown in the ordered list sequence number 5, and has the two kinase activities of lower temperature resistance pyruvate phosphate.

3. a peptide species, it has the aminoacid sequence shown in the sequence number 1 in the sequence table, and just No. 869 amino-acid substitution becomes proline(Pro).

4. a peptide species, it has the aminoacid sequence shown in the sequence number 1 in the sequence table, and just No. 885 and No. 952 amino acid are replaced as leucine and Xie Ansuan respectively.

5. the clone DNA of any one described polypeptide in the claim 1～4 of encoding.

6. a recombinant vectors wherein contains the described DNA of claim 5, can express the polypeptide with the two kinase activities of lower temperature resistance pyruvate phosphate in the host.

7. with the described DNA of claim 5 vegetable cell of transition.

Images