CN106636028A

CN106636028A - Rice protein with imidazolinone herbicide resistance activity and coding gene and application of rice protein

Info

Publication number: CN106636028A
Application number: CN201710069531.XA
Authority: CN
Inventors: 方军; 朴钟泽; 万常照; 白建江; 杨瑞芳
Original assignee: Shanghai Academy of Agricultural Sciences
Current assignee: Shanghai Academy of Agricultural Sciences
Priority date: 2017-02-08
Filing date: 2017-02-08
Publication date: 2017-05-10

Abstract

The invention discloses a rice protein with imidazolinone herbicide resistance activity and a coding gene and application of the rice protein. The rice protein is a rice acetyl hydroxy acid synthetase mutant protein, and an amino acid sequence of the rice protein is shown as SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.5 or SEQ ID NO.6. The rice protein is derived from rice genome acetyl hydroxy acid synthetase gene expression and obtained by in-vitro mutation of acetyl hydroxy acid synthetase. The amino acid sequence of the rice protein is characterized by deficiency of Ser627 locus amino acid as compared with that of wild type rice acetyl hydroxy acid synthetase. The mutant protein is resistant to imidazolinone herbicides, and plants in coding gene transformation of the rice protein have imidazolinone herbicide resistance activity.

Description

With anti-imidazolinone herbicide activity rice protein, its encoding gene and Using

Technical field

The invention belongs to herbicide resistant protein matter field, specifically relates to the paddy rice egg with anti-imidazolinone herbicide activity White matter, its encoding gene and application.

Background technology

In recent years, harm of the weeds in rice field grows in intensity.With China's expanding economy and the raising of human cost, Rice Cropping mode is from taking artificial rice transplanting progressively to the live transformation of less manpower, and the popularization of direct-seeding makes weeds in field Spread, becoming affects one of key factor of yield and quality of rice.Especially Weedy Rice, rice terrace it is constantly spontaneous, continue, Hazard rice is produced, and is the third-largest malignant weed that barnyard grass, moleplant seed are only second in rice terrace.Due to Weedy Rice and cultivated rice parent Edge is in close relations, still can distinguish Weedy Rice and cultivated rice without preferable herbicide, and plant Herbicide Resistant Rice then can be with Avoid this problem.

The endogenous antiweed Acetohydroxyacid synthase of paddy rice (acetohydroxy acid synthase, referred to as AHAS) protein is produced by ahas gene mutations, and expression antiweed AHAS protein can make plant obtain antiweed work Property, solve the weed problem in field.AHAS protein is mainly distributed in microorganism and plant, is hardly deposited in animal Suppress herbicide to person poultry harmless in, AHAS.Therefore, excavate and study the paddy rice AHAS protein tool with antiweed activity There is good novelty and economic worth.

At present, cultivate anti-AHAS to suppress the method for herbicide paddy rice is to carry out chemistry as material with rice paddy seed or plant Or physical mutagenesis, carry out genome mutation in plant body.This method obtains the test period length of Herbicide Resistant Rice and (is more than 3 years), obtain that new mutant is difficult, be mutated the replacement that result is that the single base change of nucleotides causes single amino acids, easily with The mutation that Jing is present repeats, and chance of success is little, and generally mutation success rate is less than hundred a ten thousandths, and many laboratories are surpassed Crossing the research and development of 5 years cannot obtain new mutant.

The content of the invention

It is an object of the invention to provide rice protein, its encoding gene with anti-imidazolinone herbicide activity And application, it is paddy rice AHAS muteins, and the protein has the activity of anti-imidazolinone herbicide, its coding base Because the plant for converting has the activity of anti-imidazolinone herbicide.

Different from the acid substitution mutations body for being obtained by vivo mutations at present, the present invention adopts amino acid deletion plan Slightly, by being mutated outside genosome, paddy rice AHAS protein is transformed, obtains paddy rice AHAS muteins so as to removed with anti- Careless agent activity.

In order to achieve the above object, the present invention provides following technical scheme：

Rice protein with anti-imidazolinone herbicide activity, it is paddy rice AHAS muteins, its ammonia Base acid sequence is as shown in SEQ ID No.1, SEQ ID No.2, SEQ ID No.5 or SEQ ID No.6.

Anti- imidazolone type of its amino acid sequence of the present invention as shown in SEQ ID No.1 or SEQ ID No.2 is removed The protein of careless agent activity is precursor full length protein, and after the precursor full length protein erasure signal peptide mature protein is become, Corresponding with amino acid sequence SEQ ID No.1, SEQ ID No.2, its mature amino acid sequence is respectively such as SEQ ID No.5, SEQ Shown in ID No.6.

Gene with anti-imidazolinone herbicide activity, it is that coding is described has anti-imidazolinone herbicide The nucleotide sequence of the rice protein of activity.

Further, the nucleotide sequence such as SEQ ID No.3, SEQ ID No.4, SEQ ID No.7 or SEQ ID Shown in No.8.

Rice protein with anti-imidazolinone herbicide activity of the present invention is used to cultivate anti-imidazolone type Herbicide plant.

A kind of method for obtaining anti-imidazolinone herbicide plant, including by the paddy rice AHAS muteins Encoding gene be transformed into plant, make plant produce with antiweed activity paddy rice Acetohydroxyacid synthase mutation Body protein.

Further, the nucleotide sequence of the encoding gene such as SEQ ID No.3, SEQ ID No.4, SEQ ID No.7 Or shown in SEQ ID No.8.

Further, the imidazolinone herbicide is not imazethapyr, imazamox or imazapic, but not It is limited to this several imidazolinone herbicide.

Further, the plant is paddy rice, corn or cotton, but is not limited to this several plant.

The protein of antiweed activity of the present invention, the protein is through engineered in paddy rice (Jing rice or long-grained nonglutinous rice) AHAS muteins, come from the expression of ahas1 genes in its genome, by paddy rice AHAS by external mutation, obtain The characteristics of AHAS muteins, its amino acid sequence is：Compared with wild rice AHAS protein, Ser627 positions are lacked Point amino acid, the mutant protein confrontation imidazolinone herbicide has resistance.

The encoding gene of the AHAS muteins that the present invention is obtained is transformed into plant, and table is carried out in plant Reach, make in plant containing AHAS muteins, can make plant that there is the activity of anti-imidazolinone herbicide.

The nucleotide sequence of encoding gene of the AHAS muteins of the present invention is not limited to paddy rice endogenous sequence, can be with It is the artificial synthesized nucleotide sequence for expressing the protein.

In paddy rice, corn or cotton express paddy rice AHAS mutant, obtain transgenic paddy rice R-AJM6 and R-AIM6, Transgenic corns M-AJM6 and M-AIM6, transgene cotton C-AJM6 and C-AIM6, in genetically modified plants and wild-type plant seedling Phase sprays anti-imidazolinone herbicide, and wild-type plant plant is all dead, and genetically modified plants all survive.Illustrate expression originally The plant of invention paddy rice AHAS mutant has the activity of anti-imidazolinone herbicide.

Compared with prior art, the present invention has the advantages that：

1) present invention adopts amino acid deletion strategy, and to amino acid rite-directed mutagenesis is carried out, can to deleting amino acid after it is prominent Variant proteins carry out external antiweed determination of activity, provide to develop more new antiweed AHAS muteins Basis, with simple to operate, screening process is simple, strong innovation the advantages of, excavated the endogenous anti-herbicide gene of paddy rice, increase The species of anti-herbicide gene

2) AHAS muteins of the invention are obtained by external mutation, and external mutation has designability By force, the incomparable advantage of vivo mutations such as short, strong innovation of test period, obtained the mutant time less than 2 months, meanwhile, Designed by specific mutation, it is to avoid repeat with AHAS muteins are reported.

3) after the encoding gene of AHSA muteins of the present invention is expressed, the AHAS total length muteins of acquisition With the activity that AHAS no signal peptide Ripening Mutant protein is respectively provided with anti-imidazolinone herbicide.

Description of the drawings

Fig. 1 is the AHAS protein s DS-PAGE electrophoretograms in the embodiment of the present invention 1 after purification；

Wherein, swimming lane 1：The protein of AJM6S after purification；Swimming lane 2：AIM6S protein after purification；Swimming lane 3：After purification AJWS protein；Swimming lane 4：AIWS protein after purification；Mk:Protein Marker, molecular weight is marked on the diagram.

Fig. 2 and Fig. 3 is that AJM6S and AIM6S and its control AJWS and AIWS are removed to imidazolone type in the embodiment of the present invention 2 The resistance curve map of careless agent (imazethapyr and imazamox).

Specific embodiment

Below in conjunction with specific embodiment, the invention will be further described.

In following examples, if no special instructions for《Molecular Cloning:A Laboratory guide》(Science Press, 2002) is remembered Support method.

Primer synthesizes and be sequenced student on commission's work Shanghai bioengineering limited company to be carried out, and reagent, bacterial strain, carrier etc. are real Test articles for use to buy from Hangzhou biotech company of Sigma.

Imidazoles cigarette acetic acid herbicide is " beans agree " that Shandong first reaches company, and diluted concentration is 200 times of dilution.Methyl miaow grass Cigarette herbicide is " hundred ridges lead to " of BASF Aktiengesellschaft, and diluted concentration is 1000 times of dilution.Imazamox herbicide is U.S. " the golden beans " of cyanamide company of state, diluted concentration is 100 times of dilution.

Embodiment 1 obtains the endogenous acetolactate synthestase mutein of paddy rice (Jing rice and long-grained nonglutinous rice), including following step Suddenly：

1. the ahas genes for deleting Ser627 amino acid are built using Jing rice

(1) by the method for many Polymerization chain reactions (PCR), ahas gene ajw (NCBI are amplified from Jing rice genomes Sequence number：NC_029257.1).

Wherein, entering the performing PCR primer that uses of reaction is：

RA1F：5'atggctacgaccgccgcggc；

RA1R：5'ttaatacacagtcctgccatcacca.

PCR reaction systems are：The μ l of 2 × exo+ polymerase premixed liquid 50, the μ l of primer RA1F (10 μM) 4, primer RA1R (10 μM) 4 μ l, the μ l of wild rice genomic DNA 1 and the μ l of deionized water 41, cumulative volume is 100 μ l.

PCR programs：A) 95 DEG C, 10 minutes；B) 95 DEG C, 40 seconds；C) 52 DEG C, 40 seconds；D) 68 DEG C, 2 minutes；E) circulation step 2-4,25 times；F) 68 DEG C, 10 minutes；G) 16 DEG C, preserve.

(2) separate, be purified into PCR primer, i.e. ajw genetic fragments by agarose electrophoresis, PCR primer is carried out into T clones (pGEMT, Promega companies of the U.S.), obtains T-AJW.

(3) with T-AJW as template, using primer OsA627F and OsA627R, enter performing PCR reaction, PCR primer is carried out into T Clone, obtains the mutant clone T-AJM6 for deleting Ser627 amino acid.

Wherein, in PCR reaction systems and program, removing template and outer other conditions of primer are identical with step 1 (1).

The particular sequence of primer is：

OsA627F：5'gcatgtgctgcctatgatcccagggggcgcattcaaggacatg；

OsA627R：5'catgtccttgaatgcccccctgggatcataggcagcacatgc.

Wherein, as shown in SEQ ID NO.3, amino acid sequence is as shown in SEQ ID NO.1 for the DNA sequence dna of AJM6.

2. the ahas genes for deleting Ser627 amino acid are built using long-grained nonglutinous rice：

Using the primer and experiment that are used when building the ahas genes for deleting Ser627 amino acid with above-mentioned utilization Jing rice Process.By PCR method, ahas gene aiw (NCBI sequences are amplified from long-grained nonglutinous rice genome using primer RA1F and RA1R Number：EU548084), T-AIW is obtained, then with T-AIW as template, using primer OsA627F and OsA627R, enters performing PCR reaction, PCR primer is carried out into T clones, the mutant clone T-AIM6 for deleting Ser627 amino acid is obtained, wherein, the DNA sequence dna of AIM6 As shown in SEQ ID NO.4, amino acid sequence is as shown in SEQ ID NO.2.

3. express, purify the AHAS mature proteins of erasure signal peptide

(1) T-AJW obtained with above-mentioned steps 1 (2) enters performing PCR anti-as template using primer OsABam and OsAEco Should, PCR primer is carried out into T clones, obtain the Jing rice clone T-AJWS of erasure signal peptide.

Wherein, in PCR reaction systems and program, removing template and outer other conditions of primer are identical with above-mentioned steps 1 (1), draw The particular sequence of thing is：

OsABam：5'ggatccacgccgctccggccgtgggggcc；

OsAEco：5'gaattcttaatacatagtcctgccatcac.

(2) protein expression vector is built, AJWS is cloned into pGEX4T2, the AHAS for obtaining expression no signal peptide is wild The carrier 4T2-AJWS of type protein.

(3) the carrier 4T2-RA1WS of the AHAS wild-type proteins of expression no signal peptide is proceeded into expression bacterial strain Rosetta In, centrifugation is obtained supernatant by GST posts by culture, harvesting, ultrasonic disruption cell, cleans GST posts, and wash-out is obtained Ripe Jing rice wild-type protein AJWS.

With with above-mentioned steps 3 (1)-(3) same method, with T-AIW as template, using primer OsABam and OsAEco, Enter performing PCR reaction, PCR primer is carried out into T clones, build protein expression vector, obtain the long-grained nonglutinous rice wild type of no signal peptide The carrier 4T2-AIWS of AHAS protein, then expressed by expressing bacterial strain Rosetta, obtain the ripe long-grained nonglutinous rice of no signal peptide Wild-type protein AIWS.

With with the same method of above-mentioned steps 3 (1) -2 (3), with T-AJM6 as template, using primer OsABam and OsAEco, enters performing PCR reaction, and PCR primer is carried out into T clones, builds protein expression vector, obtains no signal peptide and deletion The carrier 4T2-AJM6S of the expression maturation Jing rice AHAS muteins of Ser627 amino acid, then by expressing bacterial strain Rosetta is expressed, and is obtained no signal peptide and is deleted the ripe Jing rice AHAS muteins of Ser627 amino acid AJM6S, as shown in SEQ ID NO.5, DNA sequence dna is as shown in SEQ ID NO.7 for its amino acid sequence.

With with above-mentioned steps 3 (1)-(3) same method, with T-AIM6 as template, using primer OsABam and OsAEco, Enter performing PCR reaction, PCR primer is carried out into T clones, build protein expression vector, obtain no signal peptide and delete Ser627 ammonia The carrier 4T2-AIM6S of the ripe long-grained nonglutinous rice AHAS muteins of expression of base acid, then carry out table by expressing bacterial strain Rosetta Reach, obtain no signal peptide and delete the ripe long-grained nonglutinous rice AHAS mutein AIM6S of Ser627 amino acid, its amino acid sequence As shown in SEQ ID NO.6, DNA sequence dna is as shown in SEQ ID NO.8.

AHAS muteins AJM6S, AIM6S of wild-type protein AJWS, AIWS of acquisition and maturation are carried out SDS-PAGE electrophoresis detections, obtain protein adhesive, and protein adhesive is referring to Fig. 1.

As seen from Figure 1, the AHAS protein in swimming lane 1 to swimming lane 4 has obvious band in 90kDa positions, this position with The theoretical size of protein is consistent, meanwhile, the protein band accounts for major portion, explanation in the protein band of whole swimming lane Obtain highly purified AHAS protein.

Embodiment 2 detects the resistance of AHAS mutant protein confrontation herbicides

Variant proteins AJM6S for being obtained with embodiment 1 respectively and AIM6S, wild-type protein AJWS and AIWS are as inspection Main body is surveyed, water is blank process, with different cultivars herbicide (imidazoles cigarette acetic acid and methoxy miaow grass of the concentration at 0-100 μM Cigarette) it is influence factor, carry out antiweed biologicall test.

Solution reaction system is configured in 2ml centrifuge tubes：450 μ l measure buffer solutions, 10 μ l AHAS protein (AJM6S, AIM6S, AJWS or AIWS, 0.5 μ g/ μ l), 40 μ l influence factor solution (water be blank process, herbicide for affect because Element), the μ l of cumulative volume 500.

AHAS determination of activity buffer solutions：100mM Sodium Pyruvates, 10mM magnesium chlorides, 1mM TPP, 50 μM of FAD, 50mM phosphoric acid Salt, pH7.4.

Response procedures：Centrifuge tube containing different affecting factors reaction system is placed in into 37 DEG C to react 1 hour；20 μ l30% Sulfuric acid terminating reaction, 60 DEG C are reacted 0.5 hour, add the alpha-Naphthols of 250 μ l 1.7%, and 0.17% creatine, 60 DEG C of colour developings 0.5 are little When.200 μ l are taken out from centrifuge tube, solution of the detection containing different affecting factors is in 525nm absorbances.

Testing result：With herbicide concentration as the wild-type protein AJWS process of 0 (influence factor solution is water control) 525nm absorbances be 100% active, calculate the activity of wild type and mutein under the conditions of different affecting factors Ratio, draws curve map, as a result referring to Fig. 2-Fig. 3.

From Fig. 2-Fig. 3, with the increase of imidazolinone herbicide, the activity of wild-type protein AJWS, AIWS It is reduced to 0；And the activity of mutein AJM6S, AIM6S that 100% is still remained close in high concentration herbicide is active.

Therefore, mutein AJM6S and AIM6S of the invention have resistance to imidazolinone herbicide.

Conversion and expression of the paddy rice ahas mutant genes of embodiment 3 in plant

1. the Expression element of AHAS muteins is built

In order to express AHAS muteins in unifacial leaf and dicotyledon, with (the letter of the carriers of pCambia 1300 1300) it is called carrier, with capsicum mottle virus 35S (abbreviation 35S) promoters and terminator AHAS Expression elements is built, 35S is opened Mover and terminator DNA sequence dna are shown in http://www.snapgene.com/resources/plasmid_files/plant_ Vectors/pCAMBIA1300/, comprises the following steps that：

(1) with embodiment 1 obtain T-AJM6 as template, using primer RABam2 and RASac, by PCR and BamH1/ Sac1 double digestions, obtain ahas mutant fragments AJM6T.

Wherein, primer sequence is：

RABam2：5'gtgggatccatggctacgaccgccgcggc；

RASac：5'gtggagctcttaatacatagtcctgccatcac.

(2) it is double by PCR and BamH1/Hind3 using primer 35S-Hind and 35S-Bam with 1300 carriers as template Digestion, obtains promoter fragment 35S；

Wherein, primer sequence is：

35S-Hind：5'gcgaagcttcatggagtcaaagattcaaa；

35S-Bam：5'gtgggatccagtcccccgtgttctct ccaaatgaa.

(3) with 1300 carriers as template, using primer ter-Sac and ter-Kpn, by the double enzymes of PCR and BamH1/Hind3 Cut, obtain and terminate sub-piece ter；

Wherein, primer sequence is：

ter-Sac：5’gtggagctcagtagatgccgaccg gatctgt；

ter-Kpn：5'cagggtacccgccgaattaattcggggga.

(4) HindIII/SacI double digestions are carried out 1300, obtain fragment 1300HS, with 35S and AJM6T be attached, Clone, obtains 1300-35S-AJM6.

(5) 1300-35S-AJM6 is carried out into SacI/KpnI double digestions, is connected with ter, clones, obtain expression Jing rice AHAS The carrier 1300-AJM6 of mutant.

In the same manner, with T-AIM6 as template, by with the present embodiment step 1 (1)-(5) same steps and primer, expressed The carrier 1300-AIM6 of Jing rice AHAS mutant.

(6) the carrier 1300-AJM6 and carrier 1300-AIM6 of acquisition are proceeded to respectively in agrobacterium strains LBA4044, is obtained The Plant transformation Agrobacterium of AHAS muteins must be expressed.

2. the plant of expression AHAS muteins is obtained

The preparation method of genetically modified plants is existing mature technology, and genetically modified plants preparation commission Shanghai is rich to pray biotechnology Co., Ltd is carried out.

Carrier 1300-AJM6 and 1300-AIM6 are proceeded to respectively in paddy rice, corn and cotton using agrobacterium-mediated transformation, Obtain transgenic paddy rice R-AJM6 and R-AIM6, transgenic corns M-AJM6 and M-AIM6, transgene cotton C-AJM6 and C- AIM6。

In genetically modified plants the transgenic paddy rice R-AJM6 and R-AIM6, the transgenic corns M-AJM6 and M- that are obtained The seedling stage spray herbicide of AIM6, transgene cotton C-AJM6 and C-AIM6 and correspondence wild-type plant, after ten days, statistics is dead Rate, as a result referring to table 1.

Table 1

As shown in Table 1, wild-type plant plant is all dead, and genetically modified plants all survive, and illustrates the expression present invention The plant of paddy rice AHAS muteins there is anti-imidazolinone herbicide activity.

<110>Academy of Agricultural Sciences, Shanghai City

<120>Rice protein with anti-imidazolinone herbicide activity, its encoding gene and application

<130> 1711037

<160> 8

<170> PatentIn version 3.5

<210> SEQ ID NO.1

<211> 643

<212> PRT

<213>Paddy rice（Oryza sativa L. spp. japanica）

<400> 1

Met Ala Thr Thr Ala Ala Ala Ala Ala Ala Ala Leu Ser Ala Ala Ala

1 5 10 15

Thr Ala Lys Thr Gly Arg Lys Asn His Gln Arg His His Val Leu Pro

20 25 30

Ala Arg Gly Arg Val Gly Ala Ala Ala Val Arg Cys Ser Ala Val Ser

35 40 45

Pro Val Thr Pro Pro Ser Pro Ala Pro Pro Ala Thr Pro Leu Arg Pro

50 55 60

Trp Gly Pro Ala Glu Pro Arg Lys Gly Ala Asp Ile Leu Val Glu Ala

65 70 75 80

Leu Glu Arg Cys Gly Val Ser Asp Val Phe Ala Tyr Pro Gly Gly Ala

85 90 95

Ser Met Glu Ile His Gln Ala Leu Thr Arg Ser Pro Val Ile Thr Asn

100 105 110

His Leu Phe Arg His Glu Gln Gly Glu Ala Phe Ala Ala Ser Gly Tyr

115 120 125

Ala Arg Ala Ser Gly Arg Val Gly Val Cys Val Ala Thr Ser Gly Pro

130 135 140

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

145 150 155 160

Val Pro Met Val Ala Ile Thr Gly Gln Val Pro Arg Arg Met Ile Gly

165 170 175

Thr Asp Ala Phe Gln Glu Thr Pro Ile Val Glu Val Thr Arg Ser Ile

180 185 190

Thr Lys His Asn Tyr Leu Val Leu Asp Val Glu Asp Ile Pro Arg Val

195 200 205

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

210 215 220

Leu Val Asp Ile Pro Lys Asp Ile Gln Gln Gln Met Ala Val Pro Val

225 230 235 240

Trp Asp Thr Ser Met Asn Leu Pro Gly Tyr Ile Ala Arg Leu Pro Lys

245 250 255

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

260 265 270

Ser Arg Arg Pro Ile Leu Tyr Val Gly Gly Gly Cys Ser Ala Ser Gly

275 280 285

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

290 295 300

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

305 310 315 320

Arg Met Leu Gly Met His Gly Thr Val Tyr Ala Asn Tyr Ala Val Asp

325 330 335

Lys Ala Asp Leu Leu Leu Ala Phe Gly Val Arg Phe Asp Asp Arg Val

340 345 350

Thr Gly Lys Ile Glu Ala Phe Ala Ser Arg Ala Lys Ile Val His Ile

355 360 365

Asp Ile Asp Pro Ala Glu Ile Gly Lys Asn Lys Gln Pro His Val Ser

370 375 380

Ile Cys Ala Asp Val Lys Leu Ala Leu Gln Gly Leu Asn Ala Leu Leu

385 390 395 400

Gln Gln Ser Thr Thr Lys Thr Ser Ser Asp Phe Ser Ala Trp His Asn

405 410 415

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

420 425 430

Gly Glu Glu Ile Pro Pro Gln Tyr Ala Ile Gln Val Leu Asp Glu Leu

435 440 445

Thr Lys Gly Glu Ala Ile Ile Ala Thr Gly Val Gly Gln His Gln Met

450 455 460

Trp Ala Ala Gln Tyr Tyr Thr Tyr Lys Arg Pro Arg Gln Trp Leu Ser

465 470 475 480

Ser Ala Gly Leu Gly Ala Met Gly Phe Gly Leu Pro Ala Ala Ala Gly

485 490 495

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

500 505 510

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

515 520 525

Asn Leu Pro Val Lys Val Met Val Leu Asn Asn Gln His Leu Gly Met

530 535 540

Val Val Gln Trp Glu Asp Arg Phe Tyr Lys Ala Asn Arg Ala His Thr

545 550 555 560

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

565 570 575

Thr Ile Ala Lys Gly Phe Asn Ile Pro Ala Val Arg Val Thr Lys Lys

580 585 590

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

595 600 605

Tyr Leu Leu Asp Ile Ile Val Pro His Gln Glu His Val Leu Pro Met

610 615 620

Ile Pro Gly Gly Ala Phe Lys Asp Met Ile Leu Asp Gly Asp Gly Arg

625 630 635 640

Thr Val Tyr

<210> SEQ ID NO.2

<211> 643

<212> PRT

<213>Paddy rice（Oryza sativa L. spp. japanica）

<400> 2

Met Ala Thr Thr Ala Ala Ala Ala Ala Ala Thr Leu Ser Ala Ala Ala

1 5 10 15

Thr Ala Lys Thr Gly Arg Lys Asn His Gln Arg His His Val Leu Pro

20 25 30

Ala Arg Gly Arg Val Gly Ala Ala Ala Val Arg Cys Ser Ala Val Ser

35 40 45

Pro Val Thr Pro Pro Ser Pro Ala Pro Pro Ala Thr Pro Leu Arg Pro

50 55 60

Trp Gly Pro Ala Glu Pro Arg Lys Gly Ala Asp Ile Leu Val Glu Ala

65 70 75 80

Leu Glu Arg Cys Gly Val Ser Asp Val Phe Ala Tyr Pro Gly Gly Ala

85 90 95

Ser Met Glu Ile His Gln Ala Leu Thr Arg Ser Pro Val Ile Thr Asn

100 105 110

His Leu Phe Arg His Glu Gln Gly Glu Ala Phe Ala Ala Ser Gly Tyr

115 120 125

Ala Arg Ala Ser Gly Arg Val Gly Val Cys Val Ala Thr Ser Gly Pro

130 135 140

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

145 150 155 160

Val Pro Met Val Ala Ile Thr Gly Gln Val Pro Arg Arg Met Ile Gly

165 170 175

Thr Asp Ala Phe Gln Glu Thr Pro Ile Val Glu Val Thr Arg Ser Ile

180 185 190

Thr Lys His Asn Tyr Leu Val Leu Asp Val Glu Asp Ile Pro Arg Val

195 200 205

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

210 215 220

Leu Val Asp Ile Pro Lys Asp Ile Gln Gln Gln Met Ala Val Pro Val

225 230 235 240

Trp Asp Thr Ser Met Asn Leu Pro Gly Tyr Ile Ala Arg Leu Pro Lys

245 250 255

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

260 265 270

Ser Arg Arg Pro Ile Leu Tyr Val Gly Gly Gly Cys Ser Ala Ser Gly

275 280 285

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

290 295 300

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

305 310 315 320

Arg Met Leu Gly Met His Gly Thr Val Tyr Ala Asn Tyr Ala Val Asp

325 330 335

Lys Ala Asp Leu Leu Leu Ala Phe Gly Val Arg Phe Asp Asp Arg Val

340 345 350

Thr Gly Lys Ile Glu Ala Phe Ala Ser Arg Ala Lys Ile Val His Ile

355 360 365

Asp Ile Asp Pro Ala Glu Ile Gly Lys Asn Lys Gln Pro His Val Ser

370 375 380

Ile Cys Ala Asp Val Lys Leu Ala Leu Gln Gly Leu Asn Ala Leu Leu

385 390 395 400

Asp Gln Ser Thr Thr Lys Thr Ser Ser Asp Phe Ser Ala Trp His Asn

405 410 415

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

420 425 430

Gly Glu Glu Ile Pro Pro Gln Tyr Ala Ile Gln Val Leu Asp Glu Leu

435 440 445

Thr Lys Gly Glu Ala Ile Ile Ala Thr Gly Val Gly Gln His Gln Met

450 455 460

Trp Ala Ala Gln Tyr Tyr Thr Tyr Lys Arg Pro Arg Gln Trp Leu Ser

465 470 475 480

Ser Ala Gly Leu Gly Ala Met Gly Phe Gly Leu Pro Ala Ala Ala Gly

485 490 495

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

500 505 510

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

515 520 525

Asn Leu Pro Val Lys Val Met Val Leu Asn Asn Gln His Leu Gly Met

530 535 540

Val Val Gln Trp Glu Asp Arg Phe Tyr Lys Ala Asn Arg Ala His Thr

545 550 555 560

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

565 570 575

Thr Ile Ala Lys Gly Phe Asn Ile Pro Ala Val Arg Val Thr Lys Lys

580 585 590

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

595 600 605

Tyr Leu Leu Asp Ile Ile Val Pro His Gln Glu His Val Leu Pro Met

610 615 620

Ile Pro Gly Gly Ala Phe Lys Asp Met Ile Leu Asp Gly Asp Gly Arg

625 630 635 640

Thr Val Tyr

<210> SEQ ID NO.3

<211> 1932

<212> DNA

<213>Paddy rice（Oryza sativa L. spp. japanica）

<400> 3

atggctacga ccgccgcggc cgcggccgcc gccctgtccg ccgccgcgac ggccaagacc 60

ggccgtaaga accaccagcg acaccacgtc cttcccgctc gaggccgggt gggggcggcg 120

gcggtcaggt gctcggcggt gtccccggtc accccgccgt ccccggcgcc gccggccacg 180

ccgctccggc cgtgggggcc ggccgagccc cgcaagggcg cggacatcct cgtggaggcg 240

ctggagcggt gcggcgtcag cgacgtgttc gcctacccgg gcggcgcgtc catggagatc 300

caccaggcgc tgacgcgctc cccggtcatc accaaccacc tcttccgcca cgagcagggc 360

gaggcgttcg cggcgtccgg gtacgcgcgc gcgtccggcc gcgtcggggt ctgcgtcgcc 420

acctccggcc ccggggcaac caacctcgtg tccgcgctcg ccgacgcgct gctcgactcc 480

gtcccgatgg tcgccatcac gggccaggtc ccccgccgca tgatcggcac cgacgccttc 540

caggagacgc ccatagtcga ggtcacccgc tccatcacca agcacaatta ccttgtcctt 600

gatgtggagg acatcccccg cgtcatacag gaagccttct tcctcgcgtc ctcgggccgt 660

cctggcccgg tgctggtcga catccccaag gacatccagc agcagatggc cgtgccggtc 720

tgggacacct cgatgaatct accagggtac atcgcacgcc tgcccaagcc acccgcgaca 780

gaattgcttg agcaggtctt gcgtctggtt ggcgagtcac ggcgcccgat tctctatgtc 840

ggtggtggct gctctgcatc tggtgacgaa ttgcgctggt ttgttgagct gactggtatc 900

ccagttacaa ccactctgat gggcctcggc aatttcccca gtgacgaccc gttgtccctg 960

cgcatgcttg ggatgcatgg cacggtgtac gcaaattatg ccgtggataa ggctgacctg 1020

ttgcttgcgt ttggtgtgcg gtttgatgat cgtgtgacag ggaaaattga ggcttttgca 1080

agcagggcca agattgtgca cattgacatt gatccagcag agattggaaa gaacaagcaa 1140

ccacatgtgt caatttgcgc agatgttaag cttgctttac agggcttgaa tgctctgcta 1200

caacagagca caacaaagac aagttctgat tttagtgcat ggcacaatga gttggaccag 1260

cagaagaggg agtttcctct ggggtacaaa acttttggtg aagagatccc accgcaatat 1320

gccattcagg tgctggatga gctgacgaaa ggtgaggcaa tcatcgctac tggtgttggg 1380

cagcaccaga tgtgggcggc acaatattac acctacaagc ggccacggca gtggctgtct 1440

tcggctggtc tgggcgcaat gggatttggg ctgcctgctg cagctggtgc ttctgtggct 1500

aacccaggtg tcacagttgt tgatattgat ggggatggta gcttcctcat gaacattcag 1560

gagctggcat tgatccgcat tgagaacctc cctgtgaagg tgatggtgtt gaacaaccaa 1620

catttgggta tggtggtgca atgggaggat aggttttaca aggcgaatag ggcgcataca 1680

tacttgggca acccggaatg tgagagcgag atatatccag attttgtgac tattgctaag 1740

gggttcaata ttcctgcagt ccgtgtaaca aagaagagtg aagtccgtgc cgccatcaag 1800

aagatgctcg agactccagg gccatacttg ttggatatca tcgtcccgca ccaggagcat 1860

gtgctgccta tgatcccagg gggcgcattc aaggacatga tcctggatgg tgatggcagg 1920

actgtgtatt aa 1932

<210> SEQ ID NO.4

<211> 1932

<212> DNA

<213>Paddy rice（Oryza sativa L. spp. japanica）

<400> 4

atggctacga ccgccgcggc cgcggccgcc accttgtccg ccgccgcgac ggccaagacc 60

ggccgtaaga accaccagcg acaccacgtc cttcccgctc gaggccgggt gggggcggcg 120

gcggtcaggt gctcggcggt gtccccggtc accccgccgt ccccggcgcc gccggccacg 180

ccgctccggc cgtgggggcc ggccgagccc cgcaagggcg cggacatcct cgtggaggcg 240

ctggagcggt gcggcgtcag cgacgtgttc gcctacccgg gcggcgcgtc catggagatc 300

caccaggcgc tgacgcgctc cccggtcatc accaaccacc tcttccgcca cgagcagggc 360

gaggcgttcg cggcgtccgg gtacgcgcgc gcgtccggcc gcgtcggggt ctgcgtcgcc 420

acctccggcc ccggggcaac caacctcgtg tccgcgctcg ccgacgcgct gctcgactcc 480

gtcccgatgg tcgccatcac gggccaggtc ccccgccgca tgatcggcac cgacgccttc 540

caggagacgc ccatagtcga ggtcacccgc tccatcacca agcacaatta ccttgtcctt 600

gatgtggagg acatcccccg cgtcatacag gaagccttct tcctcgcgtc ctcgggccgt 660

cctggcccgg tgctggtcga catccccaag gacatccagc agcagatggc tgtgccagtc 720

tgggacacct cgatgaatct accggggtac attgcacgcc tgcccaagcc acccgcgaca 780

gaattgcttg agcaggtctt gcgtctggtt ggcgagtcac ggcgcccgat tctctatgtc 840

ggtggtggct gctctgcatc tggtgatgaa ttgcgccggt ttgttgagct gaccggcatc 900

ccagttacaa ccactctgat gggcctcggc aatttcccca gtgatgatcc gttgtccctg 960

cgcatgcttg ggatgcatgg cacggtgtac gcaaattatg cggtggataa ggctgacctg 1020

ttgcttgcat ttggcgtgcg gtttgatgat cgtgtgacag ggaaaattga ggcttttgca 1080

agcagggcca agattgtgca cattgacatt gatccagcgg agattggaaa gaacaagcaa 1140

ccacatgtgt caatttgcgc agatgttaag cttgctttac agggcttgaa tgctctgcta 1200

gaccagagca caacaaagac aagttctgat tttagtgcat ggcacaatga gttggaccag 1260

cagaagaggg agtttcctct ggggtacaag acttttggtg aagagatccc accgcaatat 1320

gctattcagg tgctggatga gctgacgaaa ggggaggcaa tcatcgctac tggtgttgga 1380

cagcaccaga tgtgggcggc acaatattac acctacaagc ggccacggca gtggctgtct 1440

tcggctggtc tgggcgcaat gggatttggg ctgcctgctg cagctggtgc ttctgtggct 1500

aacccaggtg tcacagttgt tgatattgat ggggatggta gcttcctcat gaacattcag 1560

gagttggcat tgatccgcat tgagaacctc ccggtgaagg tgatggtgtt gaacaaccaa 1620

catttgggta tggttgtgca atgggaggat aggttttaca aggcaaatag ggcgcataca 1680

tacttgggca acccagaatg tgagagcgag atatatccag attttgtgac tattgctaaa 1740

gggttcaata ttcctgcagt ccgtgtaaca aagaagagtg aagtccgtgc cgccatcaag 1800

aagatgctcg agaccccagg gccatacttg ttggatatca tcgtcccaca ccaggagcat 1860

gtgctgccta tgatcccagg gggcgcattc aaggacatga tcctggatgg tgatggcagg 1920

actgtgtatt aa 1932

<210> SEQ ID NO.5

<211> 583

<212> PRT

<213>Paddy rice（Oryza sativa L. spp. japanica）

<400> 5

Pro Leu Arg Pro Trp Gly Pro Ala Glu Pro Arg Lys Gly Ala Asp Ile

1 5 10 15

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

20 25 30

Pro Gly Gly Ala Ser Met Glu Ile His Gln Ala Leu Thr Arg Ser Pro

35 40 45

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

50 55 60

Ala Ser Gly Tyr Ala Arg Ala Ser Gly Arg Val Gly Val Cys Val Ala

65 70 75 80

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

85 90 95

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

100 105 110

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

115 120 125

Thr Arg Ser Ile Thr Lys His Asn Tyr Leu Val Leu Asp Val Glu Asp

130 135 140

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

145 150 155 160

Pro Gly Pro Val Leu Val Asp Ile Pro Lys Asp Ile Gln Gln Gln Met

165 170 175

Ala Val Pro Val Trp Asp Thr Ser Met Asn Leu Pro Gly Tyr Ile Ala

180 185 190

Arg Leu Pro Lys Pro Pro Ala Thr Glu Leu Leu Glu Gln Val Leu Arg

195 200 205

Leu Val Gly Glu Ser Arg Arg Pro Ile Leu Tyr Val Gly Gly Gly Cys

210 215 220

Ser Ala Ser Gly Asp Glu Leu Arg Trp Phe Val Glu Leu Thr Gly Ile

225 230 235 240

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

245 250 255

Pro Leu Ser Leu Arg Met Leu Gly Met His Gly Thr Val Tyr Ala Asn

260 265 270

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

275 280 285

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

290 295 300

Ile Val His Ile Asp Ile Asp Pro Ala Glu Ile Gly Lys Asn Lys Gln

305 310 315 320

Pro His Val Ser Ile Cys Ala Asp Val Lys Leu Ala Leu Gln Gly Leu

325 330 335

Asn Ala Leu Leu Gln Gln Ser Thr Thr Lys Thr Ser Ser Asp Phe Ser

340 345 350

Ala Trp His Asn Glu Leu Asp Gln Gln Lys Arg Glu Phe Pro Leu Gly

355 360 365

Tyr Lys Thr Phe Gly Glu Glu Ile Pro Pro Gln Tyr Ala Ile Gln Val

370 375 380

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

385 390 395 400

Gln His Gln Met Trp Ala Ala Gln Tyr Tyr Thr Tyr Lys Arg Pro Arg

405 410 415

Gln Trp Leu Ser Ser Ala Gly Leu Gly Ala Met Gly Phe Gly Leu Pro

420 425 430

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

435 440 445

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

450 455 460

Ile Arg Ile Glu Asn Leu Pro Val Lys Val Met Val Leu Asn Asn Gln

465 470 475 480

His Leu Gly Met Val Val Gln Trp Glu Asp Arg Phe Tyr Lys Ala Asn

485 490 495

Arg Ala His Thr Tyr Leu Gly Asn Pro Glu Cys Glu Ser Glu Ile Tyr

500 505 510

Pro Asp Phe Val Thr Ile Ala Lys Gly Phe Asn Ile Pro Ala Val Arg

515 520 525

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

530 535 540

Thr Pro Gly Pro Tyr Leu Leu Asp Ile Ile Val Pro His Gln Glu His

545 550 555 560

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

565 570 575

Gly Asp Gly Arg Thr Val Tyr

580

<210> SEQ ID NO.6

<211> 583

<212> PRT

<213>Paddy rice（Oryza sativa L. spp. japanica）

<400> 6

Pro Leu Arg Pro Trp Gly Pro Ala Glu Pro Arg Lys Gly Ala Asp Ile

1 5 10 15

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

20 25 30

Pro Gly Gly Ala Ser Met Glu Ile His Gln Ala Leu Thr Arg Ser Pro

35 40 45

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

50 55 60

Ala Ser Gly Tyr Ala Arg Ala Ser Gly Arg Val Gly Val Cys Val Ala

65 70 75 80

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

85 90 95

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

100 105 110

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

115 120 125

Thr Arg Ser Ile Thr Lys His Asn Tyr Leu Val Leu Asp Val Glu Asp

130 135 140

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

145 150 155 160

Pro Gly Pro Val Leu Val Asp Ile Pro Lys Asp Ile Gln Gln Gln Met

165 170 175

Ala Val Pro Val Trp Asp Thr Ser Met Asn Leu Pro Gly Tyr Ile Ala

180 185 190

Arg Leu Pro Lys Pro Pro Ala Thr Glu Leu Leu Glu Gln Val Leu Arg

195 200 205

Leu Val Gly Glu Ser Arg Arg Pro Ile Leu Tyr Val Gly Gly Gly Cys

210 215 220

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

225 230 235 240

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

245 250 255

Pro Leu Ser Leu Arg Met Leu Gly Met His Gly Thr Val Tyr Ala Asn

260 265 270

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

275 280 285

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

290 295 300

Ile Val His Ile Asp Ile Asp Pro Ala Glu Ile Gly Lys Asn Lys Gln

305 310 315 320

Pro His Val Ser Ile Cys Ala Asp Val Lys Leu Ala Leu Gln Gly Leu

325 330 335

Asn Ala Leu Leu Asp Gln Ser Thr Thr Lys Thr Ser Ser Asp Phe Ser

340 345 350

Ala Trp His Asn Glu Leu Asp Gln Gln Lys Arg Glu Phe Pro Leu Gly

355 360 365

Tyr Lys Thr Phe Gly Glu Glu Ile Pro Pro Gln Tyr Ala Ile Gln Val

370 375 380

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

385 390 395 400

Gln His Gln Met Trp Ala Ala Gln Tyr Tyr Thr Tyr Lys Arg Pro Arg

405 410 415

Gln Trp Leu Ser Ser Ala Gly Leu Gly Ala Met Gly Phe Gly Leu Pro

420 425 430

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

435 440 445

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

450 455 460

Ile Arg Ile Glu Asn Leu Pro Val Lys Val Met Val Leu Asn Asn Gln

465 470 475 480

His Leu Gly Met Val Val Gln Trp Glu Asp Arg Phe Tyr Lys Ala Asn

485 490 495

Arg Ala His Thr Tyr Leu Gly Asn Pro Glu Cys Glu Ser Glu Ile Tyr

500 505 510

Pro Asp Phe Val Thr Ile Ala Lys Gly Phe Asn Ile Pro Ala Val Arg

515 520 525

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

530 535 540

Thr Pro Gly Pro Tyr Leu Leu Asp Ile Ile Val Pro His Gln Glu His

545 550 555 560

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

565 570 575

Gly Asp Gly Arg Thr Val Tyr

58 580

<210> SEQ ID NO.7

<211> 1752

<212> DNA

<213>Paddy rice（Oryza sativa L. spp. japanica）

<400> 7

ccgctccggc cgtgggggcc ggccgagccc cgcaagggcg cggacatcct cgtggaggcg 60

ctggagcggt gcggcgtcag cgacgtgttc gcctacccgg gcggcgcgtc catggagatc 120

caccaggcgc tgacgcgctc cccggtcatc accaaccacc tcttccgcca cgagcagggc 180

gaggcgttcg cggcgtccgg gtacgcgcgc gcgtccggcc gcgtcggggt ctgcgtcgcc 240

acctccggcc ccggggcaac caacctcgtg tccgcgctcg ccgacgcgct gctcgactcc 300

gtcccgatgg tcgccatcac gggccaggtc ccccgccgca tgatcggcac cgacgccttc 360

caggagacgc ccatagtcga ggtcacccgc tccatcacca agcacaatta ccttgtcctt 420

gatgtggagg acatcccccg cgtcatacag gaagccttct tcctcgcgtc ctcgggccgt 480

cctggcccgg tgctggtcga catccccaag gacatccagc agcagatggc cgtgccggtc 540

tgggacacct cgatgaatct accagggtac atcgcacgcc tgcccaagcc acccgcgaca 600

gaattgcttg agcaggtctt gcgtctggtt ggcgagtcac ggcgcccgat tctctatgtc 660

ggtggtggct gctctgcatc tggtgacgaa ttgcgctggt ttgttgagct gactggtatc 720

ccagttacaa ccactctgat gggcctcggc aatttcccca gtgacgaccc gttgtccctg 780

cgcatgcttg ggatgcatgg cacggtgtac gcaaattatg ccgtggataa ggctgacctg 840

ttgcttgcgt ttggtgtgcg gtttgatgat cgtgtgacag ggaaaattga ggcttttgca 900

agcagggcca agattgtgca cattgacatt gatccagcag agattggaaa gaacaagcaa 960

ccacatgtgt caatttgcgc agatgttaag cttgctttac agggcttgaa tgctctgcta 1020

caacagagca caacaaagac aagttctgat tttagtgcat ggcacaatga gttggaccag 1080

cagaagaggg agtttcctct ggggtacaaa acttttggtg aagagatccc accgcaatat 1140

gccattcagg tgctggatga gctgacgaaa ggtgaggcaa tcatcgctac tggtgttggg 1200

cagcaccaga tgtgggcggc acaatattac acctacaagc ggccacggca gtggctgtct 1260

tcggctggtc tgggcgcaat gggatttggg ctgcctgctg cagctggtgc ttctgtggct 1320

aacccaggtg tcacagttgt tgatattgat ggggatggta gcttcctcat gaacattcag 1380

gagctggcat tgatccgcat tgagaacctc cctgtgaagg tgatggtgtt gaacaaccaa 1440

catttgggta tggtggtgca atgggaggat aggttttaca aggcgaatag ggcgcataca 1500

tacttgggca acccggaatg tgagagcgag atatatccag attttgtgac tattgctaag 1560

gggttcaata ttcctgcagt ccgtgtaaca aagaagagtg aagtccgtgc cgccatcaag 1620

aagatgctcg agactccagg gccatacttg ttggatatca tcgtcccgca ccaggagcat 1680

gtgctgccta tgatcccagg gggcgcattc aaggacatga tcctggatgg tgatggcagg 1740

actgtgtatt aa 1752

<210> SEQ ID NO.8

<211> 1752

<212> DNA

<213>Paddy rice（Oryza sativa L. spp. japanica）

<400> 8

ccgctccggc cgtgggggcc ggccgagccc cgcaagggcg cggacatcct cgtggaggcg 60

ctggagcggt gcggcgtcag cgacgtgttc gcctacccgg gcggcgcgtc catggagatc 120

caccaggcgc tgacgcgctc cccggtcatc accaaccacc tcttccgcca cgagcagggc 180

gaggcgttcg cggcgtccgg gtacgcgcgc gcgtccggcc gcgtcggggt ctgcgtcgcc 240

acctccggcc ccggggcaac caacctcgtg tccgcgctcg ccgacgcgct gctcgactcc 300

gtcccgatgg tcgccatcac gggccaggtc ccccgccgca tgatcggcac cgacgccttc 360

caggagacgc ccatagtcga ggtcacccgc tccatcacca agcacaatta ccttgtcctt 420

gatgtggagg acatcccccg cgtcatacag gaagccttct tcctcgcgtc ctcgggccgt 480

cctggcccgg tgctggtcga catccccaag gacatccagc agcagatggc tgtgccagtc 540

tgggacacct cgatgaatct accggggtac attgcacgcc tgcccaagcc acccgcgaca 600

gaattgcttg agcaggtctt gcgtctggtt ggcgagtcac ggcgcccgat tctctatgtc 660

ggtggtggct gctctgcatc tggtgatgaa ttgcgccggt ttgttgagct gaccggcatc 720

ccagttacaa ccactctgat gggcctcggc aatttcccca gtgatgatcc gttgtccctg 780

cgcatgcttg ggatgcatgg cacggtgtac gcaaattatg cggtggataa ggctgacctg 840

ttgcttgcat ttggcgtgcg gtttgatgat cgtgtgacag ggaaaattga ggcttttgca 900

agcagggcca agattgtgca cattgacatt gatccagcgg agattggaaa gaacaagcaa 960

ccacatgtgt caatttgcgc agatgttaag cttgctttac agggcttgaa tgctctgcta 1020

gaccagagca caacaaagac aagttctgat tttagtgcat ggcacaatga gttggaccag 1080

cagaagaggg agtttcctct ggggtacaag acttttggtg aagagatccc accgcaatat 1140

gctattcagg tgctggatga gctgacgaaa ggggaggcaa tcatcgctac tggtgttgga 1200

cagcaccaga tgtgggcggc acaatattac acctacaagc ggccacggca gtggctgtct 1260

tcggctggtc tgggcgcaat gggatttggg ctgcctgctg cagctggtgc ttctgtggct 1320

aacccaggtg tcacagttgt tgatattgat ggggatggta gcttcctcat gaacattcag 1380

gagttggcat tgatccgcat tgagaacctc ccggtgaagg tgatggtgtt gaacaaccaa 1440

catttgggta tggttgtgca atgggaggat aggttttaca aggcaaatag ggcgcataca 1500

tacttgggca acccagaatg tgagagcgag atatatccag attttgtgac tattgctaaa 1560

gggttcaata ttcctgcagt ccgtgtaaca aagaagagtg aagtccgtgc cgccatcaag 1620

aagatgctcg agaccccagg gccatacttg ttggatatca tcgtcccaca ccaggagcat 1680

gtgctgccta tgatcccagg gggcgcattc aaggacatga tcctggatgg tgatggcagg 1740

actgtgtatt aa 1752

Claims

1. there is the rice protein of anti-imidazolinone herbicide activity, it is the mutant of paddy rice Acetohydroxyacid synthase Protein, its amino acid sequence is as shown in SEQ ID No.1, SEQ ID No.2, SEQ ID No.5 or SEQ ID No.6.

2. there is the gene of anti-imidazolinone herbicide activity, it is the core for encoding rice protein described in claim 1 Nucleotide sequence.

3. as claimed in claim 2 with the gene of anti-imidazolinone herbicide activity, it is characterised in that the nucleosides Acid sequence is as shown in SEQ ID No.3, SEQ ID No.4, SEQ ID No.7 or SEQ ID No.8.

4. the rice protein as claimed in claim 1 with anti-imidazolinone herbicide activity is used to cultivate anti-imidazoline Type herbicides plant.

5. a kind of method for obtaining anti-imidazolinone herbicide plant, including by the volume of rice protein described in claim 1 Code genetic transformation makes plant produce the paddy rice acetohydroxy acid synthesis with anti-imidazolinone herbicide activity in plant The mutein of enzyme.

6. the method for obtaining anti-imidazolinone herbicide plant according to claim 5, it is characterised in that the coding base The nucleotide sequence of cause is as shown in SEQ ID No.3, SEQ ID No.4, SEQ ID No.7 or SEQ ID No.8.

7. the method for obtaining anti-imidazolinone herbicide plant according to claim 5, it is characterised in that the imidazoline Type herbicides are at least one in imazethapyr, imazamox and imazapic.

8. the method that anti-imidazolinone herbicide plant is obtained according to any one of claim 5-7, it is characterised in that institute Plant is stated for paddy rice, corn or cotton.