CN1844393A - Resistance gene Pi37 against rice blast and use thereof - Google Patents

Abstract

The invention discloses a section of nucleotide sequence and amino acid sequence, coded by the nucleotide sequence, of a new resistant gene Pi37 of rice blast and its application. The gene attribute to nonTIR-NBS-LRR resistant gene family and it is a component expression gene. The invention also relates to the application of using the gene to converse rice or other plants in order that cultivate disease-resistant variety, and the gene sequence yield labeled molecule in breeding.

Description

Resistance gene Pi 37 against rice blast and application thereof

Technical field

The present invention relates to the genetically engineered field, be specifically related to a kind of clone and application thereof of resistance gene Pi 37 against rice blast.

Background technology

Plant usually is subjected to the infringement of multiple pathogen in the process of growth, plant then takes multiple defence policies to protect self, avoids being subjected to it to attack.A most important defense mechanism is exactly the defence answering system that can discern the existence of obligate pathogenic microorganism and start self in the plant.Plant is mediated by disease-resistant gene the identification of pathogenic bacteria.Therefore, the analysis of disease-resistant gene product structure and research are the bases of understanding plant disease-resistant mechanism, also have important directive significance for the prevention and the control of Plant diseases.

So far, from 3 kinds of monocotyledonss and 5 kinds of dicotyledonss, separated more than 40 disease-resistant gene.To the structure of these disease-resistant genes and discovering of product, though host plant difference, the pathogen of institute's antagonism also has fungi, bacterium, differences such as virus and nematode, but the structure of disease-resistant gene and product have many common constitutional featuress, there is rich leucine tumor-necrosis factor glycoproteins (leucine-rich repeat as the C-end, LRR), there is nucleotide binding site (nucleotide binding site in the N-end, NBS), and leucine zipper (leucine zipper, LZ), coiled coil structural domain (coiled-coil, CC), and membrane spaning domain (transmembrane domain, TM), protein kinase (protein kinase, PK), and fruit bat Toll albumen and Mammals interleukin-1 receptor (Toll andinterleukin-1 receptor, TIR) etc.According to they coded proteic constitutional featuress, disease-resistant gene can be divided into 7 classes (Hammond-Kosack ﹠amp; Jones, 1997; Dangl ﹠amp; Jones 2001; Iyer ﹠amp; McCouch2004).

The first kind, toxin reduction enzyme disease-resistant gene.As corn disease-resistant gene Hm1, it is the 1st plant disease resistance genes that quilt is cloned, and it is responsible for the resistance to fungi Cochliobolus carbonum microspecies 1.Hm1 coding separate the HC toxin that toxenzyme can the passivation pathogenic fungi be produced, and the HC toxin is the virulence factor that fungi C.carbonum microspecies 1 produce, it determines this germ can only infect some genotype of corn (Johal etc., 1992).Second class, NBS-LRR class disease-resistant gene.The nearly N end of their encoded protein is NBS, and nearly C end then is made up of LRR.As RPS2 (Bent etc., 1994), RPM1 (Grant etc., 1995), I2 (Simon etc., 1998); RPP5 (Parker etc., 1997), N (Dodd etc., 2001), L6 (Lawrence etc., 1995), Mla1 (Zhou etc., 2001), Mla6 (Halterman etc., 2001); Disease-resistant gene of paddy rice such as Xa1 (Yoshimura etc., 1998), Pib (Wang etc., 1999), Pita (Bryan etc., 2000) etc.The 3rd class, PK class disease-resistant gene.As tomato Pto gene, its product is one and is positioned at intracellular serine-threonine protein kinase enzyme, do not have LRR structural domain (Martin etc., 1993).The 4th class, LRR-TM class disease-resistant gene.Gene C f-2 (Dixon etc., 1996), Cf-4 (Thomas etc., 1997), Cf-5 (Dixon etc., 1998), the Cf-9 (Jones etc., 1994) of the different physiological strains of the anti-leaf mold of tomato, and the gene Hs1 of the anti-Cyst nematode of beet ^Pro-I(Cai etc., 1997) etc.The 5th class, LRR-TM-PK class disease-resistant gene is representative with rice bacterial blight resistance gene Xa21 (Song etc., 1995).The 6th class is representative with the RPW8 of Arabidopis thaliana, and its encoded protein only contains complete CC and NBS structural domain (Xiao etc., 2001).The 7th class is representative with the xa5 gene of paddy rice, and its encoded protein is a transcription factor (TFIIA γ) (Iyer ﹠amp; McCouch, 2004).

The disease-resistant proteic disease-resistant gene of coding NBS-LRR class is a class disease-resistant gene maximum in the plant disease resistance genes, constructional feature according to the disease-resistant protein N terminal of NBS-LRR class, this genoid can be divided into two big class TIR-NBS-LRR (TNL) and CC-NBS-LRR (CNL) (Meyers etc. again, Pan etc., 2000; Cannon etc., 2002; Richly etc., 2002).TNL class disease-resistant gene is mainly found in dicotyledons, does not also find (Bai etc., 2002 so far in the monocotyledons genome; Meyers etc., 2002).The disease-resistant gene that in monocotyledons, identifies the at present disease-resistant albumen of CNL class of mainly encoding, also there is a large amount of CNL class disease-resistant genes in the dicotyledons, comparatively speaking, the CNL class disease-resistant gene in the monocotyledons is than more rich and varied (Cannon etc., 2002) in the dicotyledons.

Studies show that NBS, CC, TIR structural domain may participate in signal conduction (Hammond-Kosack ﹠amp; Jones 1997), although this class R albumen does not have the intrinsic kinase activity, NBS can activate kinases or G albumen, NBS has in conjunction with ATP or GTP and hydrolytic enzyme activities (Traut, 1994).The signal conduction (Jones, 1994) in the disease-resistant defense response of TIR structure possibility involved in plant downstream.Nearest evidence shows that the L family diversity of flax selects to also occur in the TIR zone, and this zone forms specificity (Luck etc., 2000) with corresponding LRR zone coevolution.The function of LRR structural domain be mainly concerned with protein-protein and with interaction (the Jones ﹠amp of part; Jones, 1996; Kajava, 1998), be product or the indirect interactional position of product (Bent, 1996 that the disease-resistant gene product is direct and the pathogenic bacteria nontoxic gene is encoded by inference; Baker etc., 1997).Jia etc. (2000) are processed as one 176 amino acid whose activated protein AVR-Pita by yeast two-hybrid proof AVR-Pita encoded protein ₁₇₆The specific exciton of microspecies is delivered to plant cytoplasm and combines with the LRD zone of Pita acceptor specifically, thus the defense response of Pita mediation in the activating cells.Av-rPita after the Ala among the Pita becomes Ser ₁₇₆Can not combine with LRD, thereby show susceptible.This result has proved that directly the LRR structural domain may be exactly the zone of pathogenic bacteria identification; Mutual " gene pairs gene " relation of making to have verified from molecular level for the first time paddy rice and Pyricularia oryzae of Pita and AvrPita.

Paddy rice is one of most important food crop in the world, and it is staple food with rice that population over half is arranged approximately.(no condition: Pyricularia grisea Sacc.) rice blast that causes is that Rice Production is endangered one of severe diseases, annually all causes serious grain loss by pathogenic fungi Magnapothe grisea Barr..From the viewpoint of Sustainable development of environment protection with agricultural, breed and the utilization of disease-resistant variety is the safe and effective procedure of control rice blast.But, because the diversity and the volatility of rice blast fungus population, people lack effective utilization of antagonism gene in addition, and antagonism mechanism lacks understanding fully, so that the susceptibleization problem of disease-resistant variety not only is not resolved, and becomes thremmatology man stubborn problem the most because of the shortage of effective anti-source gene and short-livedization of disease-resistant variety on the contrary.Therefore, excavate, identify and clone disease-resistant gene and reasonably be applied to the breeding for disease resistance plan and become the preferential major issue that solves in the agri-scientific research.

Along with molecular biological fast development, so far, have at least more than 80 rice blast master to imitate resistant gene and be in the news, wherein located for 60 by molecule.At present, also not identified the rice blast master on the 3rd karyomit(e) of paddy rice imitates the resistant gene, all identified the main resistant gene site of imitating on remaining 11 karyomit(e), and contain a plurality of rice blast resistances site on the karyomit(e) that has, and the resistant gene of some gene locus is that cluster exists.It is worthy of note in the localized resistant gene of numerous quilt successes, to have only two resistant genes of resistant gene Pib and Pita successfully to be cloned, they lay respectively on the 2nd karyomit(e) and the 12nd karyomit(e) of paddy rice.Before proposing, the present patent application also on paddy rice the 1st karyomit(e), do not clone the report of rice blast resistance gene.

Clone's disease-resistant gene is the prerequisite to the research of paddy rice resistance mechanism, discloses the molecule mechanism of rice anti-rice blast and can control and reduce the harm of Pyricularia oryzae to paddy rice better.Simultaneously,, can control and increase the disease resistance of plant artificially, widen the anti-spectrum of plant clone's the modification and the transformation of disease-resistant gene.These aspects are that employing conventional plant breeding and improving technology institute are inaccessiable.

Summary of the invention

A rice blast resistance gene that the objective of the invention is to carry among the separating clone rice varieties Q1333 and the dna fragmentation that comprises the promotor of regulating and control this gene.

Another object of the present invention provides the coded protein of above-mentioned rice blast resistance gene.

Another object of the present invention provides the above-mentioned carrier that contains above-mentioned resistant gene.

Another object of the present invention provides above-mentioned carrier transgenic plant transformed.

Another object of the present invention provides the application of above-mentioned protein in preparation resisting rice blast bacteria medicine.

Further purpose of the present invention provides molecule marker that above-mentioned resistant gene produces and rice blast is had application in the paddy rice of disease resistance in seed selection.

The present invention relates to separate and use a kind of dna fragmentation of the Pi37 of comprising gene, this fragment is given plant the caused disease of Pyricularia oryzae (Magnaporthe grisea) is produced specific disease resistance response.This invention is applicable to all plants to this pathogenic bacteria sensitivity.These plants comprise monocotyledons and dicotyledons.Wherein, described fragment perhaps is equivalent to the dna sequence dna shown in the SEQ ID NO:1 basically shown in sequence table SEQ ID NO:1, and perhaps its function is equivalent to the subfragment of sequence shown in the SEQ ID NO:1.A kind of NBS-LRR proteinoid of this dna sequence encoding, its aminoacid sequence such as SEQ ID NO:2 are listed, and structure is as shown in Figure 6.Dna fragmentation shown in the present is constitutive expression at the leaf tissue of paddy rice.

The Pi37 resistant gene coding NBS-LRR albumen that separates, clones.This albumen comprises two main structural domain: NBS and LRR zone, and wherein the NBS structural domain contains conservative kinase 1a:GGAGKS, is positioned at 222-227 amino-acid residue of this polypeptide; Kinase 2a:LLVLDDV is positioned at 297-303 amino-acid residue of this polypeptide; Kinase 3a:GSRVLVTSRR is positioned at 327-336 amino-acid residue of this polypeptide.And the 589-1290 of this a proteic C-end amino-acid residue is 25 LRR repetitions, and its leucine content is 17.0%.

The nucleotide fragments of coding NBS or LRR may have independently function in the Pi37 gene.Fragment and the reorganization of other nucleic acid fragments with coding different structure territory in the Pi37 gene can constitute mosaic gene or protein, make it to have new function.The Pi37 gene is modified or transformed, can change or increase certain function of gene.For example, the LRR zone of this gene is replaced with the structural domain of other resistant genes, maybe the NBS structural domain with this gene carries out rite-directed mutagenesis, may cause the change of the anti-spectrum of the forfeiture of gene resistance or gene.

The present invention comprises that equally the primary structure part with the Pi37 resistant gene effectively connects the upward formed mosaic gene of proper regulation sequence, and comprises the plant of this gene and the seed of this kind of plant in genome.As this gene can be natural or chimeric.For example, will comprise the fragment of this gene and the promotor of a constitutive expression and be connected, this promotor can and be expressed under any condition cytocerastic any period.The promotor of this constitutive expression comprises the promotor of cauliflower mosaic virus 35S etc.On the other hand, also promotor that can the promotor of this gene and a tissue specific expression or developmental stage is specific expressed or accurately the promotor of environmental induction be connected, these promotors are referred to as inducible promoter.Like this, the change of environment, the difference of developmental stage can change this expression of gene, and is same, also this expression of gene can be limited in some tissues, makes by this gene induced disease resistance response and obtains artificial control.Wherein envrionment conditions comprises attack, anaerobic condition and the light etc. of pathogenic bacteria, and tissue and developmental stage comprise leaf, fruit, seed and flower etc.

According to Pi37 gene order information provided by the invention (SEQ ID NO:1), those skilled in the art can easily obtain the gene that is equal to Pi36 by the following method: (1) obtains by database retrieval; (2) with the Pi37 gene fragment be genomic library or the acquisition of cDNA library of probe Screening of Rice or other plant; (3) according to Pi37 gene order information design oligonucleotides primer, from genome, mRNA and the cDNA of paddy rice or other plant, obtain with the method for pcr amplification; (4) on the basis of Pi37 gene order, obtain with the gene engineering method transformation; (5) method with chemosynthesis obtains this gene.

Rice blast resistance gene Pi37 provided by the invention has important use and is worth.One of application is that described Pi37 gene order is connected to any plant conversion carrier, with any method for transformation the Pi37 disease-resistant gene is imported paddy rice or other plant cell, can obtain the transgenosis disease-resistant variety of expressing said gene, thereby be applied to produce.Of the present invention gene constructed in plant conversion carrier, can suitably modify described gene or its regulating and controlling sequence, also can before its transcription initiation codon, replace the original promotor of described gene, thereby widen and strengthen the resistance of plant pathogenic bacteria with other promotor.

The Another application of resistant gene provided by the invention is to produce specific molecule marker according to described gene order information, includes but not limited to SNP (mononucleotide polymorphic), SSR (simple sequence repeats polymorphic), RFLP (restriction enzyme length is polymorphic), CAP (the cutting amplified fragments is polymorphic).Can identify the resistant gene type of paddy rice or other plant with this mark, be used for the molecular marker assisted selection breeding, thereby improve the efficiency of selection of breeding.

The present invention has following beneficial effect: change clone's disease-resistant gene over to susceptible plant, help to produce new disease-resistant plants.Particularly can be with transformation technology a plurality of disease-resistant genes that in plant, add up, and can not produce the chain problem of bad gene in the genome of following appearance in the traditional breeding technology, and can shorten breeding time.The clone of disease-resistant gene can overcome the problem that can not shift disease-resistant gene in the traditional breeding method between plant species.In addition, disease-resistant transfer-gen plant that the present invention can further provide or the above-mentioned dna fragmentation of applications exploiting obtains and corresponding seed, and with gene of the present invention or based on the recombinant chou plants transformed of this gene or the seed that obtains by this class plant.Can gene of the present invention be changed over to other plant with the mode of sexual hybridization.

Description of drawings

Fig. 1 is the map based cloning synoptic diagram of resistance gene Pi 37 against rice blast;

Fig. 2 is the high-res genetic map and the electronics physical map of resistance gene Pi 37 against rice blast;

Fig. 3 a is the PCR detected result figure of hygromycin gene of the resistance transformant of resistance gene Pi 37 against rice blast;

Fig. 3 b is the Southern hybridization figure of hygromycin gene of the resistance transformant of resistance gene Pi 37 against rice blast;

Fig. 4 is that the resistance of resistance gene Pi 37 against rice blast transformed plant is identified figure;

Fig. 5 is the gene structure figure of resistance gene Pi 37 against rice blast;

Fig. 6 is a resistance gene Pi 37 against rice blast amino acids coding peptide sequence structure iron;

Fig. 7 is that the RT-PCR of resistance gene Pi 37 against rice blast and allelotrope pi37 expression characterization thereof detects figure;

The Pi37 loci gene type that Fig. 8 identifies parent and disease plant for molecule marker FSTS1 is figure as a result.

Wherein, A represents the genetic map of Pi37 gene among Fig. 2, and map distance is centimorgan (cM); B represents the physical map of Pi37 gene, the numeral recombination event below the mark in the bracket, the numeral physical distance (kb) between the mark; C represents to make up the contig of Pi37 gene physical map, and short sea line is represented the fine BAC/PAC clone of japonica rice variety Japan that IRGSP announces, vertical line is represented mark place clone's relative position.Swimming lane 1 is molecular weight marker DL2000 among Fig. 3 a; Swimming lane 2,3 and 4 is respectively the conversion seedling T of resistant gene Pi37 ₀-1, T ₀-2 and T ₀The pcr amplification product of-3 hygromycin gene; Swimming lane 5 is the non-transformant of acceptor Q1063; Swimming lane 6 is the PCR product of the hygromycin gene of carrier pCAMBIA1300 plasmid; Swimming lane 7 is the clear water contrast.

Swimming lane 1 is the non-transformant of donor Q1333 among Fig. 3 b; Swimming lane 2 is the non-transformant of acceptor Q1063; Swimming lane 3,4 and 5 is respectively the conversion seedling T of resistant gene Pi37 ₀-1, T ₀-2 and T ₀The pcr amplification product of-3 hygromycin gene; Swimming lane 6 is molecular weight marker DL2000.

Green square box is represented the exon of resistant gene Pi37 among Fig. 5, and the square box of band oblique line is respectively 5 ' and 3 ' non-translational region.

Underlined letter is for forming 3 zones of inferring NBS among Fig. 6; The amino-acid residue that independently list the lower section is the terminal LRR of C-zone.

A is that disease-resistant variety Q1333 is before inoculation and the expression conditions of inoculation back 24h and 48h among Fig. 7; B is susceptible variety Q13, i.e. LTH is before inoculation and the expression conditions of inoculation back 24h and 48h; Actin is an internal reference.

M is the DL2000DNA standard molecular weight among Fig. 8; 1 is disease-resistant parent Q1333; 2-4 is respectively susceptible parent C101PKT, CO39 and AS20-1; 5-7 and 9-11 are susceptible individual; 8 is disease-resistant individuality; The arrow indication is flag F STS1.

Embodiment

Embodiment 1: the genetic analysis of resistance gene Pi 37 against rice blast and Primary Location

The present invention is to 3 F by the cross combination origin of japonica rice disease-resistant variety Q1333 and 3 long-grained nonglutinous rice susceptible variety AS20-1, C101PKT and CO39 ₂Colony inoculates bacterial strain CHL1159, CHL1340 and CHL1405 to the clearly demarcated non-affinity/compatible host response of parents' kind performance respectively, and the result shows this 3 F ₂Disease-resistant plant was all controlled by a pair of dominant gene with separating than all meeting 3R: 1S, infer Q1333 thus the resistance to above-mentioned 3 inoculating strains that is showed of disease plant in the colony.To these 3 F ₂The resistant gene of colony's correspondence carries out linkage analysis, and the result shows that they are controlled by same dominant gene, has therefore made up from these 3 F ₂Colony's origin, totally 85 disease-resistant individualities and 701 big mapping populations that susceptible individual is formed.

Utilize little satellite (simple sequence repeat, SSR) molecular marking technique, in conjunction with based on segregating population analytical method (bulked-sgregant analysis, BSA) recessive population analysis method (recessive-classanalysis, RCA), this goal gene has been carried out linkage analysis.180 microsatellite markers antagonism sense parents and anti-sense gene pool that utilization equidistant from 12 karyomit(e)s of paddy rice (10cM approximately at interval) is selected screen, and have obtained 2 candidate's linked marker RM128 and RM486 that are positioned on the 1st karyomit(e).The resistant gene that these 3 colonies of this presentation of results identify is different with the Pif gene.In order to confirm this result, between anti-sense parent, there are microsatellite marker RM543, RM302, RM212 and the RM319 of polymorphism to carry out linkage analysis having chosen 4 between RM128 and the RM486 again, the result shows SSR mark RM543 and the more close purpose resistance of RM319 site, is respectively 0.7cM and 1.6cM with the genetic distance of goal gene; Two other SSR mark RM302 and RM212 then with goal gene be divided into fully from.It may be another one new resistance gene on the Q1333 that this result further illustrates resistant gene that this research identifies, its called after Pi37.

Embodiment 2: the Fine Mapping of resistance gene Pi 37 against rice blast

For the Pi37 site is narrowed down in the littler genome area, utilize the reference sequences of public database order-checking kind Japan fine (Nipponbare), by bioinformatic analysis (bioinformatics analysis, BIA) method has been developed new molecule marker at target genome area, promptly new SSR and STS mark.In 8 flag F PSM1, FPSM2, FPSM3, FPSM4, FSTS1, FSTS2, FSTS3 and FSTS4 of exploitation, have only flag F PSM3 not show polymorphism; Further recombination event analysis revealed, FPSM1 detects 1 recombinant chou, FPSM2 and FSTS2 detect 2 recombinant chous, other 4 flag F PSM4, FSTS1, FSTS3 and FSTS4 then with goal gene be divided into fully from.The presentation of results of comprehensive above-mentioned linkage analysis, Pi37 (t) site is defined at the heredity zone of 0.77cM between flank mark RM543 and the FPSM1, in this zone, have 7 molecule marker RM302, FPSM4, RM212, FSTS4, S15628, FSTS1 and FSTS3 all be divided into fully with it from.

In order to make up the physical map in Pi37 site, the mark with Pi37 gene linkage is all landed on above-mentioned reference sequences by the BIA method, and made up the contig in this goal gene site thus.Can infer that by reference sequences the Pi37 site is defined in the scope of 374kb between flank mark RM543 and the FPSM1.Further analyze by BIA, confirmed that having only 4 in this zone has the resistant gene conserved structure, nucleotide binding site and rich leucine repeat (nucleotide binding site-leucine-rich repeat, candidate gene NBS-LRR).Then further this goal gene site is defined on the dna fragmentation of about 60kb (Fig. 2) thus.

Embodiment 3: the conversion of rice blast new resistance gene Pi37 and the resistance of transformant are identified

In order to determine the function of candidate resistance gene, 4 candidate gene R37-L1, R37-L2, R37-L3 and R37-L4 of above prediction have been carried out function complementation experiment respectively.(long-range PCR, LR-PCR) increased respectively 4 candidate resistance genes and be cloned into binary vector transformation system pCAMBIA1300 of technology have imported agrobacterium strains EHA105 to utilize long segment PCR.Will height susceptible variety Q1063 mature seed evoked callus on inducing culture.The EHA105 that contains the goal gene conversion carrier was cultivated 1 day for 28 ℃ at the YM nutrient agar, be collected in the MB liquid nutrient medium of the Syringylethanone that contains 100 μ mol/L.The rice callus tissue was immersed bacterium liquid 20 minutes, blot and transfer to the MB nutrient agar behind the bacterium liquid and cultivate.Shift callus to the culture medium culturing that contains the 50mg/L Totomycin, subculture was 1 time in per 14 days, subculture 2 times.Behind the resistance screening, change regeneration culture medium over to and differentiate the conversion seedling, obtained 6 strains of conversion seedling, 68 strains, 92 strains and 79 strains of R37-L1, R37-L2, R37-L3 and R37-L4 respectively.

The PCR that transformed plant has been carried out hygromycin gene identifies (Fig. 3).Detected result proof order gene fragment has imported these transformant.The resistance qualification result shows to have only candidate gene R37-L3 to show the identical resistance with resistant gene donor Q1333 under susceptible variety Q1063 genetic background, shows that resistant gene Pi37 has successfully been cloned (Fig. 4).

The structure of embodiment 4:Pi37 gene

The employing method of moving one's steps is measured the dna sequence dna of Pi37.Utilize 5 ' and 3 ' RACE reaction, obtained the full-length cDNA of Pi37 and it is checked order.Pi37 gene DNA length is 7486bp, and its full-length cDNA is 4691bp, contains the opening code-reading frame of a 3873bp, and 5 ' and 3 ' non-translational region is respectively 192bp and 626bp (Fig. 5).

The structure of embodiment 5:Pi37 resistance protein

The protein sequence of Pi37 genes encoding is shown in SEQ ID NO:2 in the sequence table.Resistant gene Pi37 1 protein polypeptide of forming by 1290 amino-acid residues of encoding, molecular weight is 120KD, iso-electric point is 6.61.Pi37 albumen belongs to NBS-LRR albumen, and conservative kinase 1a (GGAGKS) in the NBS structural domain is positioned at 222-227 amino-acid residue of this polypeptide; Kinase 2a (LLVLDDV) is positioned at 297-303 amino-acid residue of this polypeptide; Kinase 3a (GSRVLVTSRR) is positioned at 327-336 amino-acid residue of this polypeptide.And the 590-1290 of this a proteic C-end amino-acid residue is 25 LRR repetitions, and its leucine content is 17.0%.

Embodiment 6:Pi37 expression of gene specificity analysis

With RT-PCR Pi37 expression of gene pattern is analyzed.From the blade of disease-resistant variety St.No.1 and susceptible variety LHT, extract total RNA, utilize reverse transcription test kit SuperScript ^TMReverseTranscriptase II carries out the synthetic of reverse transcription cDNA article one chain.The RT-PCR primer is R37RT F:CCTTGGTGGGCTTACTTCACTTAG; R36RT R:TCCGTCAAGTGCCTGAGGTTATG.The PCR reaction is: 94 ℃ of pre-sex change 4min; Next be 35 circulations, cycling program is as follows: 94 ℃ of sex change 30sec, and 56 ℃ of annealing 30sec, 72 ℃ are extended 1min; Last 72 ℃ are extended 7min, and temperature drops to 4 ℃ and promptly finishes amplification.Experimental result shows, the RNA reverse transcription template of the leaf tissue before and after the inoculation of resistant variety and susceptible variety all can amplify special fragment, illustrate that Pi37 and allelotrope pi37 thereof all can express in leaf tissue, promptly belong to the gene (Fig. 7) of constitutive expression.

Embodiment 7: transform the application of the resistant variety of Pi37 gene generation

The Pi37 gene clone to plant conversion carrier pCAMBIA1300, is imported agrobacterium strains EHA105, use and transform susceptible rice varieties, obtained 92 strain transformed plants.The conversion that transformant is produced is (T ₁) inoculate evaluation, find wherein to have 20 to transform system's performance resistances and separate, and the resistance of transformed plant and Pi37 gene be divided into from.Explanation can be used Pi37 gene transformation paddy rice susceptible variety, is used for producing.

The application of embodiment 8:Pi37 gene order in the molecular marker assisted selection breeding

Utilize Pi37 gene order information provided by the invention can produce molecule marker, be used to identify that the range gene type that has on the Pi37 site is the plant of Pi37Pi37, Pi37pi37 and pi37pi37, can in the molecular marker assisted selection breeding process, be applied.

Embodiment 9: the resistance feature of resistant gene Pi37

At first use parent strain CHL42 and CHL43 and 78 offspring's bacterial strains thereof, the 3 cover breed combinations of being made up of 15 kinds have been carried out anti-spectrum comparative analysis.The result shows that 14 entrained resistance significant reaction that resistant gene showed of Pi37 gene and other rice varieties are different, particularly compares with the anti-spectrum of other resistant gene on the 1st karyomit(e), and difference is more obvious.Next use 5 colonies of collecting from 5 provinces of Chinese Guangdong, Fujian, Hunan, Yunnan and Jiangsu totally 537 bacterial strains carried out pathogenic mensuration, the result shows the Pi37 gene except Jiangsu, and the bacterial strain that other major part is picked up from China all has the height resistance of wide spectrum.

Resistance gene Pi 37 against rice blast and application sequence table thereof

SEQUENCE?LISTING

＜110〉Agricultural University Of South China

＜120〉resistance gene Pi 37 against rice blast and application thereof

<130>

<160>2

<170>PatentIn?version?3.2

<210>1

<211>7486

<212>DNA

＜213〉Oryza paddy rice (Orysa sativa L.)

<220>

<221>5’UTR

<222>(1)..(1343)

<220>

<221>exon

<222>(1344)..(5216)

<220>

<221>gene

<222>(1344)..(5216)

<220>

<221>3’UTR

<222>(5217)..(7486)

<400>1

gaaacagcta?tgacatgatt?acgaattcga?gctcggtacc?cggggatcct?ctagagtcga 60

ccctaccgag?tccagcaaaa?tccattgcat?atccttcatt?ttgtcgaagc?tgtgccagtc 120

catctcttca?tggtcatttc?atcatcacca?gctccccgtc?ggagctggcc?ttcattgtca 180

cttcatatct?tcatcatctc?catgtcggag?cttgtcatca?tcatcatcaa?tcagtgaagg 240

tccgaagaac?agtttgtgaa?gggccttcat?caatatcaat?gtctcatcat?caatcaaagg 300

accgaagaac?agcttgtgaa?ggtccattgt?catcatcatc?aacatctccg?aatcaatgaa 360

gagccgaaga?acgattcgtg?aaggcttcaa?ggtaccgagg?gccgaagaac?agtttgtgaa 420

ggctccgagg?tccatttcat?tgaggagggg?gtcttttctt?cttcatcatc?caagaattga 480

cacacttcga?cgacatcatc?aagactttgg?agataggcta?atatttagat?tgtgttctca 540

tgtttagttt?acctttcaaa?tgcaatgtta?ttacatacac?attgcgtttg?agtgggggtg 600

ttaagggtag?tatgtctagg?ggtattttag?atatttccca?tctctctctt?gtacaaatat 660

attttgccct?ttgggcctaa?tctaattgga?acagttcatt?ccccaatttc?tccaaattgt 720

gacacagctc?atctgaatat?tagagaaata?taaatttgcc?actaacattt?aacaatgtcc 780

cacttgtcta?ctccctccgt?caaaaaaaac?caaacctaag?acttagatgt?gatataatct 840

agtatctgtc?tagattcgtt?gtactagatt?atggcttaaa?tactagatta?agtttttttt 900

gggacggagg?gagtatgtct?aaccaaaata?acacaattga?tttgcatatt?gtgttttgcc 960

tcgcattgtg?ccgaagcaat?cttatctgtg?gcaattggta?gagttcttcc?tctcacacct 1020

tctctgcaat?acaaacgcat?ctggctctgg?taaccttgca?ctcctctctt?agtatttctt 1080

tccttcacgc?tttcttgtct?ctgcttaagt?tctctatcgt?ttggtaccat?catgcattaa 1140

ttcctttttc?ctctatttcc?cactccaaac?atttccataa?ccactcacat?acacaaaaat 1200

aaaaaaccag?acagagagaa?aaaaacttgt?tttgggagtt?ctgttgttat?cttcctcaca 1260

gaagaaaaat?ctctccatca?ccatctgttc?ttatgccctg?cttctattca?ttcacttgtt 1320

cacagctgca?gtagtgctgt?tcc?atg?gcg?gag?gtg?gtg?ttg?gct?ggc?tta?cgt 1373

Met?Ala?Glu?Val?Val?Leu?Ala?Gly?Leu?Arg

1 5 10

ttg?gca?gca?aca?cca?acc?tgt?gtg?aag?ctc?ctc?tgt?aat?gct?tcg?acg 1421

Leu?Ala?Ala?Thr?Pro?Thr?Cys?Val?Lys?Leu?Leu?Cys?Asn?Ala?Ser?Thr

15 20 25

tgc?ctt?ggc gtg?gac?atg?acg?cgt?gag?ctc?cat?gaa?ctg?gag?acc?atc 1469

Cys?Leu?Gly?Val?Asp?Met?Thr?Arg?Glu?Leu?His?Glu?Leu?Glu?Thr?Ile

30 35 40

atc?ata?cca?cag?ttc?gag?ctg?gtg?att?gaa?gca?gct?gag?aaa?gga?aac 1517

Ile?Ile?Pro?Gln?Phe?Glu?Leu?Val?Ile?Glu?Ala?Ala?Glu?Lys?Gly?Asn

45 50 55

cac?aga?gcc?aag?ctg?gac?aga?tgg?ctc?cgg?gag?ctc?aaa?caa?gcc?ttc 1565

His?Arg?Ala?Lys?Leu?Asp?Arg?Trp?Leu?Arg?Glu?Leu?Lys?Gln?Ala?Phe

60 65 70

tac?aat?gct?gag?gac?ctt?ctg?gac?gag?cat?gag?tac?aac?atc?ctt?aag 1613

Tyr?Asn?Ala?Glu?Asp?Leu?Leu?Asp?Glu?His?Glu?Tyr?Asn?Ile?Leu?Lys

75 80 85 90

cgc?aaa?gcg?aaa?cac?aag?gat?tct?cta?gtg?aaa?gat?tcc?acc?cag?gtt 1661

Arg?Lys?Ala?Lys?His?Lys?Asp?Ser?Leu?Val?Lys?Asp?Ser?Thr?Gln?Val

95 100 105

cat?gac?tct?tcc?atc?agc?aat?att?ttg?aag?cag?cct?atg?cgt?gct?gtg 1709

His?Asp?Ser?Ser?Ile?Ser?Asn?Ile?Leu?Lys?Gln?Pro?Met?Arg?Ala?Val

110 115 120

tcc?agt?agg?atg?tcc?aat?ctg?cgc?cca?gag?aac?aga?aag?ata?ctt?tgc 1757

Ser?Ser?Arg?Met?Ser?Asn?Leu?Arg?Pro?Glu?Asn?Arg?Lys?Ile?Leu?Cys

125 130 135

cag?ttg?aat?gaa?ctg?aag?acc?atg?ctg?gag?aaa?gcc?aag?gag?ttc?cgt 1805

Gln?Leu?Asn?Glu?Leu?Lys?Thr?Met?Leu?Glu?Lys?Ala?Lys?Glu?Phe?Arg

140 145 150

gag?ctg?att?cac?tta?cct?gct?ggt?aat?agt?ctt?gag?ggt?ccc?agt?gta 1853

Glu?Leu?Ile?His?Leu?Pro?Ala?Gly?Asn?Ser?Leu?Glu?Gly?Pro?Ser?Val

155 160 165 170

cca?aca?att?gtt?gtt?cct?gta?gtg?aca?tcg?cta?ttg?ccg?cca?aga?gta 1901

Pro?Thr?Ile?Val?Val?Pro?Val?Val?Thr?Ser?Leu?Leu?Pro?Pro?Arg?Val

175 180 185

ttt?ggt?cgc?aac?atg?gat?cgc?gat?cgc?ata?ata?cat?ctt?ctt?act?aag 1949

Phe?Gly?Arg?Asn?Met?Asp?Arg?Asp?Arg?Ile?Ile?His?Leu?Leu?Thr?Lys

190 195 200

cca?atg?gct?act?gtt?tct?agc?tca?gtc?ggc?tac?tct?ggt?ttg?gcc?att 1997

Pro?Met?Ala?Thr?Val?Ser?Ser?Ser?Val?Gly?Tyr?Ser?Gly?Leu?Ala?Ile

205 210 215

gtt?gcg?cat?gga?gga?gca?gga?aaa?tcc?acc?tta?gca?caa?tgt?gtt?tac 2045

Val?Ala?His?Gly?Gly?Ala?Gly?Lys?Ser?Thr?Leu?Ala?Gln?Cys?Val?Tyr

220 225 230

aat?gac?aag?agg?gta?caa?gaa?cat?ttt?gac?gtc?agg?ata?tgg?gtc?tgc 2093

Asn?Asp?Lys?Arg?Val?Gln?Glu?His?Phe?Asp?Val?Arg?Ile?Trp?Val?Cys

235 240 245 250

atc?tca?cgc?aaa?ctt?gat?gtt?cat?cgc?cat?aca?cgt?gag?att?atc?gag 2141

Ile?Ser?Arg?Lys?Leu?Asp?Val?His?Arg?His?Thr?Arg?Glu?Ile?Ile?Glu

255 260 265

tca?gca?aca?aat?gga?gag?tgc?cct?cgt?gtg?gac?aat?ctt?gat?act?ctc 2189

Ser?Ala?Thr?Asn?Gly?Glu?Cys?Pro?Arg?Val?Asp?Asn?Leu?Asp?Thr?Leu

270 275 280

cag?tgc?aga?ttg?aag?gac?ata?atg?caa?aaa?tca?gag?aaa?ttc?ctg?ctt 2237

Gln?Cys?Arg?Leu?Lys?Asp?Ile?Met?Gln?Lys?Ser?Glu?Lys?Phe?Leu?Leu

285 290 295

gtg?ttg?gat?gac?gtc?tgg?ttt?gat?gaa?tcc?gtc?aat?gag?agg?gaa?tgg 2285

Val?Leu?Asp?Asp?Val?Trp?Phe?Asp?Glu?Ser?Val?Asn?Glu?Arg?Glu?Trp

300 305 310

gac?cag?ttg?ctt?gat?cct?ctt?gtt?tct?cag?cag?gag?ggg?agc?aga?gtt 2333

Asp?Gln?Leu?Leu?Asp?Pro?Leu?Val?Ser?Gln?Gln?Glu?Gly?Ser?Arg?Val

315 320 325 330

ttg?gtg?act?tct?aga?cga?gat?gta?ctt?cca?gct?gct?ctt?cac?tgt?aag 2381

Leu?Val?Thr?Ser?Arg?Arg?Asp?Val?Leu?Pro?Ala?Ala?Leu?His?Cys?Lys

335 340 345

ggt?gtt?gtt?cat?ttg?gaa?aac?atg?gaa?gat?gcc?gag?ttc?ttg?gct?cta 2429

Gly?Val?Val?His?Leu?Glu?Asn?Met?Glu?Asp?Ala?Glu?Phe?Leu?Ala?Leu

350 355 360

ttc?aaa?tac?cat?gct?ttc?tct?gga?aca?gaa?atc?aga?aat?cca?cag?ttg 2477

Phe?Lys?Tyr?His?Ala?Phe?Ser?Gly?Thr?Glu?Ile?Arg?Asn?Pro?Gln?Leu

365 370 375

cat?gca?cgg?cta?gaa?gag?gtc?gca?gag?aag?att?gct?aaa?agg?tta?gga 2525

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

380 385 390

caa?tcg?cct?tta?gca?gca?agg?aca?gta?ggc?tcc?cag?ctg?agc?agg?aat 2573

Gln?Ser?Pro?Leu?Ala?Ala?Arg?Thr?Val?Gly?Ser?Gln?Leu?Ser?Arg?Asn

395 400 405 410

aag?gat?att?gcc?ata?tgg?aaa?agt?gct?cta?aat?att?gaa?aac?tta?agt 2621

Lys?Asp?Ile?Ala?Ile?Trp?Lys?Ser?Ala?Leu?Asn?Ile?Glu?Asn?Leu?Ser

415 420 425

gag?ccc?atg?aaa?gct?ctg?ttg?tgg?agt?tat?aat?aaa?tta?gat?tca?cgt 2669

Glu?Pro?Met?Lys?Ala?Leu?Leu?Trp?Ser?Tyr?Asn?Lys?Leu?Asp?Ser?Arg

430 435 440

ctg?cag?aga?tgt?ttt?ctg?tac?tgc?agc?tta?ttt?cca?aaa?ggt?cac?aag 2717

Leu?Gln?Arg?Cys?Phe?Leu?Tyr?Cys?Ser?Leu?Phe?Pro?Lys?Gly?His?Lys

445 450 455

tat?aaa?att?gat?gag?atg?gtt?gac?ctt?tgg?gtg?gca?gag?ggg?cta?gtt 2765

Tyr?Lys?Ile?Asp?Glu?Met?Val?Asp?Leu?Trp?Val?Ala?Glu?Gly?Leu?Val

460 465 470

gat?tca?cgc?aac?cag?ggg?gat?aag?aga?att?gaa?gat?att?ggc?agg?gat 2813

Asp?Ser?Arg?Asn?Gln?Gly?Asp?Lys?Arg?Ile?Glu?Asp?Ile?Gly?Arg?Asp

475 480 485 490

tac?ttc?aat?gag?atg?gtg?tca?gga?tct?ttc?ttt?caa?cca?gtt?tct?gaa 2861

Tyr?Phe?Asn?Glu?Met?Val?Ser?Gly?Ser?Phe?Phe?Gln?Pro?Val?Ser?Glu

495 500 505

aga?tat?atg?ggt?aca?tgg?tac?att?atg?cat?gat?ctg?ctt?cat?gat?ttg 2909

Arg?Tyr?Met?Gly?Thr?Trp?Tyr?Ile?Met?His?Asp?Leu?Leu?His?Asp?Leu

510 515 520

gca?gag?tca?ctg?act?aaa?gaa?gac?tgc?ttc?aga?tta?gaa?gat?gat?ggg 2957

Ala?Glu?Ser?Leu?Thr?Lys?Glu?Asp?Cys?Phe?Arg?Leu?Glu?Asp?Asp?Gly

525 530 535

gtg?aaa?gag?ata?cca?gcc?act?gtt?cga?cat?cta?tct?att?tgt?gtt?gac 3005

Val?Lys?Glu?Ile?Pro?Ala?Thr?Val?Arg?His?Leu?Ser?Ile?Cys?Val?Asp

540 545 550

agt?atg?aaa?ttc?cat?aag?caa?aag?atc?tgc?aag?cta?cgt?tat?tta?cgc 3053

Ser?Met?Lys?Phe?His?Lys?Gln?Lys?Ile?Cys?Lys?Leu?Arg?Tyr?Leu?Arg

555 560 565 570

act?gtt?atc?tgc?att?gac?cct?ctc?atg?gat?gac?gga?gat?gat?att?ttt 3101

Thr?Val?Ile?Cys?Ile?Asp?Pro?Leu?Met?Asp?Asp?Gly?Asp?Asp?Ile?Phe

575 580 585

aat?cag?cta?ttg?aag?aat?ctg?aag?aag?cta?cgt?gta?cta?cat?ttg?tcg 3149

Asn?Gln?Leu?Leu?Lys?Asn?Leu?Lys?Lys?Leu?Arg?Val?Leu?His?Leu?Ser

590 595 600

ttt?tac?aac?agt?agc?agc?ttg?cct?gaa?tgt?att?ggt?gag?ctg?aag?cac 3197

Phe?Tyr?Asn?Ser?Ser?Ser?Leu?Pro?Glu?Cys?Ile?Gly?Glu?Leu?Lys?His

605 610 615

ctt?cga?tat?ttg?agt?atc?atc?agc?aca?ttg?att?tct?gaa?ctg?cca?aga 3245

Leu?Arg?Tyr?Leu?Ser?Ile?Ile?Ser?Thr?Leu?Ile?Ser?Glu?Leu?Pro?Arg

620 625 630

tca?ttg?tgt?act?ctt?ttc?cac?ttg?gag?cta?ctt?cat?ttg?aat?gac?aag 3293

Ser?Leu?Cys?Thr?Leu?Phe?His?Leu?Glu?Leu?Leu?His?Leu?Asn?Asp?Lys

635 640 645 650

gtc?aag?aat?ttg?ccg?gac?aga?ctc?tgt?aat?tta?aga?aag?tta?cga?cgt 3341

Val?Lys?Asn?Leu?Pro?Asp?Arg?Leu?Cys?Asn?Leu?Arg?Lys?Leu?Arg?Arg

655 660 665

ctt?gaa?gca?tat?gat?gac?aga?aac?aga?gtg?tat?aaa?ttg?tat?aga?gca 3389

Leu?Glu?Ala?Tyr?Asp?Asp?Arg?Asn?Arg?Val?Tyr?Lys?Leu?Tyr?Arg?Ala

670 675 680

gct?ctc?cct?caa?att?cct?tac?ata?ggc?aag?cta?tct?ttg?ctg?caa?gat 3437

Ala?Leu?Pro?Gln?Ile?Pro?Tyr?Ile?Gly?Lys?Leu?Ser?Leu?Leu?Gln?Asp

685 690 695

att?gat?gga?ttt?tgc?gtg?caa?aag?cag?aag?gga?tat?gag?ttg?cga?caa 3485

Ile?Asp?Gly?Phe?Cys?Val?Gln?Lys?Gln?Lys?Gly?Tyr?Glu?Leu?Arg?Gln

700 705 710

ctg?agg?gac?atg?aac?aag?ctc?ggt?ggt?aat?ttg?cgt?gtc?gtt?aat?ctt 3533

Leu?Arg?Asp?Met?Asn?Lys?Leu?Gly?Gly?Asn?Leu?Arg?Val?Val?Asn?Leu

715 720 725 730

gaa?aat?gta?act?gga?aag?gat?gaa?gcc?tca?gag?tcg?aag?ttg?cat?cag 3581

Glu?Asn?Val?Thr?Gly?Lys?Asp?Glu?Ala?Ser?Glu?Ser?Lys?Leu?His?Gln

735 740 745

aaa?act?cat?ctt?aga?ggt?ttg?cat?ctc?tcc?tgg?aat?gat?gtg?gat?gac 3629

Lys?Thr?His?Leu?Arg?Gly?Leu?His?Leu?Ser?Trp?Asn?Asp?Val?Asp?Asp

750 755 760

atg?gat?gtt?tca?cat?ttg?gag?att?cta?gaa?ggt?ttg?agg?ccg?cca?tct 3677

Met?Asp?Val?Ser?His?Leu?Glu?Ile?Leu?Glu?Gly?Leu?Arg?Pro?Pro?Ser

765 770 775

caa?ttg?gag?gac?ctt?aca?atc?gaa?ggt?tac?aaa?tcc?acc?atg?tac?cca 3725

Gln?Leu?Glu?Asp?Leu?Thr?Ile?Glu?Gly?Tyr?Lys?Ser?Thr?Met?Tyr?Pro

780 785 790

agc?tgg?tta?ctt?gat?gga?tcc?tat?ttt?gag?aat?cta?gag?tct?ttt?acg 3773

Ser?Trp?Leu?Leu?Asp?Gly?Ser?Tyr?Phe?Glu?Asn?Leu?Glu?Ser?Phe?Thr

795 800 805 810

ctt?gct?aat?tgc?tgt?gtg?ata?gga?agc?cta?cca?ccc?aat?acc?gag?atc 3821

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

815 820 825

ttt?agg?cac?tgt?atg?aca?ctt?acc?ctt?gag?aat?gtc?ccg?aat?atg?aag 3869

Phe?Arg?His?Cys?Met?Thr?Leu?Thr?Leu?Glu?Asn?Val?Pro?Asn?Met?Lys

830 835 840

aca?cta?cct?ttt?ctt?cca?gaa?ggt?ctc?aca?agc?cta?tca?att?gag?ggg 3917

Thr?Leu?Pro?Phe?Leu?Pro?Glu?Gly?Leu?Thr?Ser?Leu?Ser?Ile?Glu?Gly

845 850 855

tgc?cca?ctg?ctt?gtg?ttc?act?act?aat?aac?gat?gaa?ccg?gaa?cac?cat 3965

Cys?Pro?Leu?Leu?Val?Phe?Thr?Thr?Asn?Asn?Asp?Glu?Pro?Glu?His?His

860 865 870

gat?tat?agg?gag?agc?atc?acg?agg?gca?aac?aac?ctg?gaa?aca?caa?ctt 4013

Asp?Tyr?Arg?Glu?Ser?Ile?Thr?Arg?Ala?Asn?Asn?Leu?Glu?Thr?Gln?Leu

875 880 885 890

gtt?ttg?ata?tgg?gaa?gcg?aat?tcc?gat?tca?gat?att?agg?agc?aca?ttg 4061

Val?Leu?Ile?Trp?Glu?Ala?Asn?Ser?Asp?Ser?Asp?Ile?Arg?Ser?Thr?Leu

895 900 905

tcg?tcc?gaa?cac?tca?tcc?atg?aag?aag?ttg?acg?gaa?ttg?atg?gat?act 4109

Ser?Ser?Glu?His?Ser?Ser?Met?Lys?Lys?Leu?Thr?Glu?Leu?Met?Asp?Thr

910 915 920

gat?atg?tcg?gga?aat?ctt?caa?acc?att?gaa?agc?gct?tta?gaa?ata?gag 4157

Asp?Met?Ser?Gly?Asn?Leu?Gln?Thr?Ile?Glu?Ser?Ala?Leu?Glu?Ile?Glu

925 930 935

aga?gat?gaa?gcc?ttg?gtt?aaa?gaa?gat?atc?atc?aaa?gta?tgg?ctc?tgt 4205

Arg?Asp?Glu?Ala?Leu?Val?Lys?Glu?Asp?Ile?Ile?Lys?Val?Trp?Leu?Cys

940 945 950

tgc?cac?gag?gag?agg?atg?aga?ttc?att?tgt?tca?agg?aag?gct?ggg?ttg 4253

Cys?His?Glu?Glu?Arg?Met?Arg?Phe?Ile?Cys?Ser?Arg?Lys?Ala?Gly?Leu

955 960 965 970

ccg?ttg?gtt?cta?cca?tct?gga?cta?tgc?gta?ctc?tct?ctt?tcc?tcc?tgt 4301

Pro?Leu?Val?Leu?Pro?Ser?Gly?Leu?Cys?Val?Leu?Ser?Leu?Ser?Ser?Cys

975 980 985

agt?att?aca?gat?gga?gct?tta?gct?att?tgc?ctt?ggt?ggg?ctt?act?tca 4349

Ser?Ile?Thr?Asp?Gly?Ala?Leu?Ala?Ile?Cys?Leu?Gly?Gly?Leu?Thr?Ser

990 995 1000

ctt?aga?aat?ttg?ttc?tta?aca?gag?att?atg?act?tta?act?acg?ctt 4394

Leu?Arg?Asn?Leu?Phe?Leu?Thr?Glu?Ile?Met?Thr?Leu?Thr?Thr?Leu

1005 1010 1015

cca?ccg?gaa?gag?gtc?ttc?caa?cat?ttg?ggc?aat?ctt?aga?tac?ttg 4439

Pro?Pro?Glu?Glu?Val?Phe?Gln?His?Leu?Gly?Asn?Leu?Arg?Tyr?Leu

1020 1025 1030

gtc?att?cgt?tcc?tgc?tgg?tgt?ttc?aga?tca?ttc?ggg?ggc?ttg?cga 4484

Val?Ile?Arg?Ser?Cys?Trp?Cys?Phe?Arg?Ser?Phe?Gly?Gly?Leu?Arg

1035 1040 1045

tct?gct?acc?tct?ctt?tca?gaa?ata?aga?tta?ttt?tcc?tgt?cct?tct 4529

Ser?Ala?Thr?Ser?Leu?Ser?Glu?Ile?Arg?Leu?Phe?Ser?Cys?Pro?Ser

1050 1055 1060

tta?cag?ttg?gca?cgt?ggc?gca?gaa?ttt?atg?caa?atg?tcc?ctt?gag 4574

Leu?Gln?Leu?Ala?Arg?Gly?Ala?Glu?Phe?Met?Gln?Met?Ser?Leu?Glu

1065 1070 1075

aag?cta?tgt?gta?tac?aac?tgt?gtg?ctt?tca?gct?gac?ttc?ttc?tgt 4619

Lys?Leu?Cys?Val?Tyr?Asn?Cys?Val?Leu?Ser?Ala?Asp?Phe?Phe?Cys

1080 1085 1090

ggt?gac?tgg?cca?cat?ctg?gat?gat?att?ctc?tta?tct?ggg?tgc?aga 4664

Gly?Asp?Trp?Pro?His?Leu?Asp?Asp?Ile?Leu?Leu?Ser?Gly?Cys?Arg

1095 1100 1105

agc?tcg?tcg?tcc?ttg?cat?gtt?ggt?gat?ctt?acc?tcc?ctc?gaa?tca 4709

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

1110 1115 1120

ttc?tca?cta?tat?cat?ttt?cca?gat?tta?tgc?acg?ctg?gaa?ggt?ttg 4754

Phe?Ser?Leu?Tyr?His?Phe?Pro?Asp?Leu?Cys?Thr?Leu?Glu?Gly?Leu

1125 1130 1135

tct?tcc?ctg?cag?ctt?cac?cac?gtg?cat?ttg?ata?gat?gtt?cca?aag 4799

Ser?Ser?Leu?Gln?Leu?His?His?Val?His?Leu?Ile?Asp?Val?Pro?Lys

1140 1145 1150

ctt?act?acc?gag?agc?atc?tca?cag?ttt?cgc?gtc?cag?cgt?tcg?ctc 4844

Leu?Thr?Thr?Glu?Ser?Ile?Ser?Gln?Phe?Arg?Val?Gln?Arg?Ser?Leu

1155 1160 1165

tat?att?agc?agt?tct?gtt?atg?ctc?aac?cac?atg?ctc?tct?gct?gaa 4889

Tyr?Ile?Ser?Ser?Ser?Val?Met?Leu?Asn?His?Met?Leu?Ser?Ala?Glu

1170 1175 1180

ggt?ttc?gtt?gtt?cca?gag?ttt?ctc?tct?ctt?gaa?agc?tgc?aag?gag 4934

Gly?Phe?Val?Val?Pro?Glu?Phe?Leu?Ser?Leu?Glu?Ser?Cys?Lys?Glu

1185 1190 1195

cca?tcc?gtt?tca?ttc?gaa?gaa?tct?gca?aac?ttc?aca?tcc?gtc?aag 4979

Pro?Ser?Val?Ser?Phe?Glu?Glu?Ser?Ala?Asn?Phe?Thr?Ser?Val?Lys

1200 1205 1210

tgc?ctg?agg?tta?tgt?aat?tgt?gaa?atg?agg?tca?cca?cca?ggg?aat 5024

Cys?Leu?Arg?Leu?Cys?Asn?Cys?Glu?Met?Arg?Ser?Pro?Pro?Gly?Asn

1215 1220 1225

atg?aag?tgc?cta?tcc?agt?cta?acg?aaa?ctc?gat?atc?tat?gat?tgc 5069

Met?Lys?Cys?Leu?Ser?Ser?Leu?Thr?Lys?Leu?Asp?Ile?Tyr?Asp?Cys

1230 1235 1240

ccc?aac?ata?tca?tct?ata?cca?gat?ctg?ccg?tcc?tcc?ctc?cag?cat 5114

Pro?Asn?Ile?Ser?Ser?Ile?Pro?Asp?Leu?Pro?Ser?Ser?Leu?Gln?His

1245 1250 1255

ata?tgt?ata?tgg?ggt?tgt?gag?ctc?ttg?aag?gag?agc?tgc?aga?gca 5159

Ile?Cys?Ile?Trp?Gly?Cys?Glu?Leu?Leu?Lys?Glu?Ser?Cys?Arg?Ala

1260 1265 1270

cct?gaa?gga?gaa?agc?tgg?cca?aag?att?gcg?cat?atc?cgc?tgg?aag 5204

Pro?Glu?Gly?Glu?Ser?Trp?Pro?Lys?Ile?Ala?His?Ile?Arg?Trp?Lys

1275 1280 1285

gaa?ttc?aga?tga?attgtgcttc?agaatcacaa?ctagagagaa?acggaaaggt 5256

Glu?Phe?Arg

1290

gaaggttaca?tcaatgcatt?gtcagaagta?ctttcttttc?tgtattattt?atttcgttaa 5316

cccatgatca?tgcaaacaca?ttatttactt?caatgtttcc?ttgtttattt?attttgaaca 5376

gacaccccaa?gattggcaca?tatacgtttc?cctgctgcca?tccaccgact?cgccgcctcc 5436

ttttattcgg?tttctctgcc?actgacggcc?catgatcgtg?tcctgttgct?taaatggagt 5496

cttgttggct?taagataatc?tctgtgtctc?ttgaagactc?aactatcttg?tatttgttag 5556

gatacgatag?gcaaccaaac?cataaaaaac?aatagatcaa?tcataaaata?tacgagattt 5616

taacgtggaa?aaccctacca?aaacaggaga?gaaaaaacca?cgggcgtcag?ccagcaaaat 5676

atctttacta?tatctggggt?gaggttacaa?cgccgcacgt?cggcttacaa?gaggtatata 5736

tatggtggaa?ccctagaagg?aatgacgatc?cggcccaagc?ctcccggcga?cgggcctccg 5796

ctccactacg?gcagaagctg?gcctttatta?agtgcagatg?aatttggatc?ataactcaac 5856

agtattttat?tatgtgatgt?gatatactat?tccagtactt?ggattttaca?cttcacacaa 5916

atatacagtt?tgcttcaaat?gtttgtctat?gtagtttgca?taaagtttgt?atggatatcg 5976

attatttgtg?taagtttctg?atgcttttat?cgcacgatag?ttcttgatag?ttctcacctc 6036

acttcgaatg?tgtccttgac?tccttgttga?gctgcccggc?agtctgacaa?aatttgatca 6096

gtccccattg?tttctgttgc?actctggcgc?ttggattgtt?tgacttagtt?ctctctctat 6156

cctcccggtt?gagtaagata?taggtggtgt?ttgaatctcc?tgaagataaa?aataaagata 6216

aaaattaaat?gtttcacaca?aaacgaggtg?gtaataacgt?gtgagtaatt?aagctttaat 6276

cattacaaac?ttgaaaaata?gattaatatg?atattttaga?acaactttca?tatagaaagt 6336

tttcacacga?aacgaaccgt?ttagctgttt?aaaaagcgtg?ccacgagtat?ccaaaagttt 6396

atccaacgtt?tgttagcaga?ttaattttga?taatagtagc?agaatgtccg?tccgttgcta 6456

ccggtggaag?cacgtgactt?cgcgcgtcac?ctacttcctc?cgtttcataa?tggaagtcat 6516

tctaacattt?cccacattta?cagtgatgtt?tctagattca?ttaacatcaa?tatgaatgtg 6576

gaaaatgcta?gaatgactta?cattgtgaaa?cggagggagt?agttgttagt?tcgcaatata 6636

attatgggaa?cataagaaaa?acttactaag?aatagtttaa?cccaatatat?cgattcttaa 6696

ttttcatcaa?tagaaagggg?catccgagta?tccgacagaa?ctgaatattg?gactctctgg 6756

ttgaatatgc?atacatgtac?tgagcactga?tccagatcaa?taaaaagacg?ttactgacac 6816

ggaaacacag?catcaggtat?gcacgaactg?aaagcataac?tctgcatgat?tgcatctttg 6876

gctaacggct?gtctatggag?tgttagacgt?cgataaattc?aacccagaga?tcattccagc 6936

ccagctatgc?atgattgcat?ctttgcctag?cctacttagt?gaccaactgc?cccctccgtt 6996

tcagattata?agatgttttt?gactttgatc?aaagtcaaac?tgtttcaaat?ttaactaaat 7056

ttatagaaaa?aaatagtaat?attctccacc?caacataaat?ttattattaa?agtacattta 7116

attattaatt?taataaaact?aatttggtaa?tgtaaatatt?actttatttc?tctataaatt 7176

tagtctaact?ttatacaatt?taactttgat?caaagtcaaa?acatcttata?acttaaaacg 7236

ggtggagata?ttgggatacg?ccccacgtag?gagcgttagg?tatgcccagg?taacggagct 7296

taccctttac?attcctcagt?tctagcatat?ccttgctgtg?ccagattggg?aaacgtcgac 7356

ctgcaggcat?gcaagcttgg?cactggccgt?cgttttacaa?cgtcgtgact?gggaaaaccc 7416

tggcgttacc?caacttaatc?gccttgcagc?acatccccct?ttcgccagct?ggcgtaatag 7476

cgaagaggcc 7486

<210>2

<211>1291

<212>NBS-LRR

＜213〉Oryza paddy rice (Orysa sativa L.)

<400>2

Met?Ala?Glu?Val?Val?Leu?Ala?Gly?Leu?Arg?Leu?Ala?Ala?Thr?Pro

5 10 15

Ile?Cys?Val?Lys?Leu?Leu?Cys?Asn?Ala?Ser?Thr?Cys?Leu?Gly?Val

20 25 30

Asp?Met?Thr?Arg?Glu?Leu?His?Glu?Leu?Glu?Thr?Ile?Ile?Ile?Pro

35 40 45

Gln?Phe?Glu?Leu?Val?Ile?Glu?Ala?Ala?Glu?Lys?Gly?Asn?His?Arg

50 55 60

Ala?Lys?Leu?Asp?Arg?Trp?Leu?Arg?Glu?Leu?Lys?Gln?Ala?Phe?Tyr

65 70 75

Asn?Ala?Glu?Asp?Leu?Leu?Asp?Glu?His?Glu?Tyr?Asn?Ile?Leu?Lys

80 85 90

Cys?Lys?Ala?Lys?His?Lys?Asp?Ser?Leu?Val?Lys?Asp?Ser?Thr?Gln

95 100 105

Val?His?Asp?Ser?Ser?Ile?Ser?Asn?Ile?Leu?Lys?Gln?Pro?Met?Arg

110 115 120

Ala?Val?Ser?Ser?Arg?Met?Ser?Asn?Leu?Arg?Pro?Glu?Asn?Arg?Lys

125 130 135

Ile?Leu?Cys?Gln?Leu?Asn?Glu?Leu?Lys?Thr?Met?Leu?Glu?Lys?Ala

140 145 150

Lys?Glu?Phe?Arg?Glu?Leu?Ile?His?Leu?Pro?Ala?Gly?Asn?Ser?Leu

155 160 165

Glu?Gly?Pro?Ser?Val?Pro?Thr?Ile?Val?Val?Pro?Val?Val?Thr?Ser

170 175 180

Leu?Leu?Pro?Pro?Arg?Val?Phe?Gly?Arg?Asn?Met?Asp?Arg?Asp?Arg

185 190 195

Ile?Ile?His?Leu?Leu?Thr?Lys?Pro?Met?Ala?Thr?Val?Ser?Ser?Ser

200 205 210

Val?Gly?Tyr?Ser?Gly?Leu?Ala?Ile?Val?Ala?His?Gly?Gly?Ala?Gly

215 220 225

Lys?Ser?Thr?Leu?Ala?Gln?Cys?Val?Tyr?Asn?Asp?Lys?Arg?Ala?Gln

230 235 240

Glu?His?Phe?Asp?Val?Arg?Met?Trp?Val?Cys?Ile?Ser?Arg?Lys?Leu

245 250 255

Asp?Val?His?Arg?His?Thr?Arg?Glu?Ile?Ile?Glu?Ser?Ala?Thr?Asn

260 265 270

Gly?Glu?Cys?Pro?Arg?Val?Asp?Asn?Leu?Asp?Thr?Leu?Gln?Cys?Arg

275 280 285

Leu?Lys?Asp?Ile?Met?Gln?Lys?Ser?Glu?Lys?Phe?Leu?Leu?Val?Leu

290 295 300

Asp?Asp?Val?Trp?Phe?Asp?Glu?Ser?Val?Asn?Glu?Arg?Glu?Trp?Asp

305 310 315

Gln?Leu?Leu?Asp?Pro?Leu?Val?Ser?Gln?Gln?Glu?Gly?Ser?Arg?Val

320 325 330

Leu?Val?Thr?Ser?Arg?Arg?Asp?Val?Leu?Pro?Ala?Ala?Leu?His?Cys

335 340 345

Lys?Asp?Val?Val?His?Leu?Glu?Asn?Met?Glu?Asp?Ala?Glu?Phe?Leu

350 355 360

Ala?Leu?Phe?Lys?Tyr?His?Ala?Phe?Ser?Gly?Thr?Glu?Ile?Arg?Asn

365 370 375

Pro?Gln?Leu?His?Ala?Arg?Leu?Glu?Glu?Val?Ala?Glu?Lys?Ile?Ala

380 385 390

Lys?Arg?Leu?Gly?Gln?Ser?Pro?Leu?Ala?Ala?Arg?Thr?Val?Gly?Ser

395 400 405

Gln?Leu?Ser?Arg?Asn?Lys?Asp?Ile?Ala?Ile?Trp?Lys?Ser?Ala?Leu

410 415 420

Asn?Ile?Glu?Asn?Leu?Ser?Glu?Pro?Met?Lys?Ala?Leu?Leu?Trp?Ser

425 430 435

Tyr?Asn?Lys?Leu?Asp?Ser?Arg?Leu?Gln?Arg?Cys?Phe?Leu?Tyr?Cys

440 445 450

Ser?Leu?Phe?Pro?Lys?Gly?His?Lys?Tyr?Lys?Ile?Asp?Glu?Met?Val

455 460 465

Asp?Leu?Trp?Val?Ala?Glu?Gly?Leu?Val?Asp?Ser?Arg?Asn?Gln?Gly

470 475 480

Asp?Lys?Arg?Ile?Glu?Asp?Ile?Gly?Arg?Asp?Tyr?Phe?Asn?Glu?Met

485 490 495

Val?Ser?Gly?Ser?Phe?Phe?Gln?Pro?Val?Ser?Glu?Arg?Tyr?Met?Gly

500 505 510

Thr?Trp?Tyr?Ile?Met?His?Asp?Leu?Leu?His?Asp?Leu?Ala?Glu?Ser

515 520 525

Leu?Thr?Lys?Glu?Asp?Cys?Phe?Arg?Leu?Glu?Asp?Asp?Gly?Val?Lys

530 535 540

Glu?Ile?Pro?Ala?Thr?Val?Arg?His?Leu?Ser?Ile?Cys?Val?Asp?Ser

545 550 555

Met?Lys?Phe?His?Lys?Gln?Lys?Ile?Cys?Lys?Leu?Arg?Tyr?Leu?Arg

560 565 570

Thr?Val?Ile?Cys?Ile?Asp?Pro?Leu?Met?Asp?Asp?Gly?Asp?Asp?Ile

575 580 585

Phe?Asn?Gln?Leu?Leu?Lys?Asn?Leu?Lys?Lys?Leu?Arg?Val?Leu?His

590 595 600

Leu?Ser?Phe?Tyr?Asn?Ser?Ser?Ser?Leu?Pro?Glu?Cys?Ile?Gly?Glu

605 610 615

Leu?Lys?His?Leu?Arg?Tyr?Leu?Ser?Ile?Ile?Ser?Thr?Leu?Ile?Ser

620 625 630

Glu?Leu?Pro?Arg?Ser?Leu?Cys?Thr?Leu?Phe?His?Leu?Glu?Leu?Leu

635 640 645

His?Leu?Asn?Asp?Lys?Val?Lys?Asn?Leu?Pro?Asp?Arg?Leu?Cys?Asn

650 655 660

Leu?Arg?Lys?Leu?Arg?Arg?Leu?Glu?Ala?Tyr?Asp?Asp?Arg?Asn?Arg

665 670 675

Met?Tyr?Lys?Leu?Tyr?Arg?Ala?Ala?Leu?Pro?Gln?Ile?Pro?Tyr?Ile

680 685 690

Gly?Lys?Leu?Ser?Leu?Leu?Gln?Asp?Ile?Asp?Gly?Phe?Cys?Val?Gln

695 700 705

Lys?Gln?Lys?Gly?Tyr?Glu?Leu?Arg?Gln?Leu?Arg?Asp?Met?Asn?Lys

710 715 720

Leu?Gly?Gly?Asn?Leu?Arg?Val?Val?Asn?Leu?Glu?Asn?Val?Thr?Gly

725 730 735

Lys?Asp?Glu?Ala?Ser?Glu?Ser?Lys?Leu?His?Gln?Lys?Thr?His?Leu

740 745 750

Arg?Gly?Leu?His?Leu?Ser?Trp?Asn?Asp?Val?Asp?Asp?Met?Asp?Val

755 760 765

Ser?His?Leu?Glu?Ile?Leu?Glu?Gly?Leu?Arg?Pro?Pro?Ser?Gln?Leu

770 775 780

Glu?Asp?Leu?Thr?Ile?Glu?Gly?Tyr?Lys?Ser?Thr?Met?Tyr?Pro?Ser

785 790 795

Trp?Leu?Leu?Asp?Gly?Ser?Tyr?Phe?Glu?Asn?Leu?Glu?Ser?Phe?Thr

800 805 810

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

815 820 825

Ile?Phe?Arg?His?Cys?Met?Thr?Leu?Thr?Leu?Glu?Asn?Val?Pro?Asn

830 835 840

Met?Lys?Thr?Leu?Pro?Phe?Leu?Pro?Glu?Gly?Leu?Thr?Ser?Leu?Ser

845 850 855

Ile?Glu?Gly?Cys?Pro?Leu?Leu?Val?Phe?Thr?Thr?Asn?Asn?Asp?Glu

860 865 870

Leu?Glu?His?His?Asp?Tyr?Arg?Glu?Ser?Ile?Thr?Arg?Ala?Asn?Asn

875 880 885

Leu?Glu?Thr?Gln?Leu?Val?Leu?Ile?Trp?Glu?Ala?Asn?Ser?Asp?Ser

890 895 900

Asp?Ile?Arg?Ser?Thr?Leu?Ser?Ser?Glu?His?Ser?Ser?Met?Lys?Lys

905 910 915

Leu?Thr?Glu?Leu?Met?Asp?Thr?Asp?Met?Ser?Gly?Asn?Leu?Gln?Thr

920 925 930

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

935 940 945

Glu?Asp?Ile?Ile?Lys?Val?Trp?Leu?Cys?Cys?His?Glu?Glu?Arg?Met

950 955 960

Arg?Phe?Ile?Tyr?Ser?Arg?Lys?Ala?Gly?Leu?Pro?Leu?Val?Leu?Pro

965 970 975

Ser?Gly?Leu?Cys?Val?Leu?Ser?Leu?Ser?Ser?Cys?Ser?Ile?Thr?Asp

980 985 990

Gly?Ala?Leu?Ala?Ile?Cys?Leu?Gly?Gly?Leu?Thr?Ser?Leu?Arg?Asn

995 1000 1005

Leu?Phe?Leu?Thr?Glu?Ile?Met?Thr?Leu?Thr?Thr?Leu?Pro?Pro?Glu

1010 1015 1020

Glu?Val?Phe?Gln?His?Leu?Gly?Asn?Leu?Arg?Tyr?Leu?Val?Ile?Arg

1025 1030 1035

Ser?Cys?Trp?Cys?Leu?Arg?Ser?Phe?Gly?Gly?Leu?Arg?Ser?Ala?Thr

1040 1045 1050

Ser?Leu?Ser?Glu?Ile?Arg?Leu?Phe?Ser?Cys?Pro?Ser?Leu?Gln?Leu

1055 1060 1065

Ala?Arg?Gly?Ala?Glu?Phe?Met?Gln?Met?Ser?Leu?Glu?Lys?Leu?Cys

1070 1075 1080

Val?Tyr?Asn?Cys?Val?Leu?Ser?Ala?Asp?Phe?Phe?Cys?Gly?Asp?Trp

1085 1090 1095

Pro?His?Leu?Asp?Asp?Ile?Leu?Leu?Ser?Gly?Cys?Arg?Ser?Ser?Ser

1100 1105 1110

Ser?Leu?His?Val?Gly?Asp?Leu?Thr?Ser?Leu?Glu?Ser?Phe?Ser?Leu

1115 1120 1125

Tyr?His?Phe?Pro?Asp?Leu?Cys?Thr?Leu?Glu?Gly?Leu?Ser?Ser?Leu

1130 1135 1140

Gln?Leu?His?His?Val?His?Leu?Ile?Asp?Val?Pro?Lys?Leu?Thr?Thr

1145 1150 1155

Glu?Ser?Ile?Ser?Gln?Phe?Arg?Val?Gln?Arg?Ser?Leu?Tyr?Ile?Ser

1160 1165 1170

Ser?Ser?Val?Met?Leu?Asn?His?Met?Leu?Ser?Ala?Glu?Gly?Phe?Val

1175 1180 1185

Val?Pro?Glu?Phe?Leu?Ser?Leu?Glu?Ser?Cys?Lys?Glu?Pro?Ser?Val

1190 1195 1200

Ser?Phe?Glu?Glu?Ser?Ala?Asn?Phe?Thr?Ser?Val?Lys?Cys?Leu?Arg

1205 1210 12L5

Leu?Cys?Asn?Cys?Glu?Met?Arg?Ser?Pro?Pro?Gly?Asn?Met?Lys?Cys

1220 1225 1230

Leu?Ser?Ser?Leu?Thr?Lys?Leu?Asp?Ile?Tyr?Asp?Cys?Pro?Asn?Ile

1235 1240 1245

Ser?Ser?Ile?Pro?Asp?Leu?Pro?Ser?Ser?Leu?Gln?His?Ile?Cys?Ile

1250 1255 1260

Trp?Gly?Cys?Glu?Leu?Leu?Lys?Glu?Ser?Cys?Arg?Ala?Pro?Glu?Gly

1265 1270 1275

Glu?Ser?Trp?Pro?Lys?Ile?Ala?His?Ile?Arg?Trp?Lys?Glu?Phe?Arg

1280 1285 1290

ter

Claims (9)

1, a kind of resistance gene Pi 37 against rice blast encoded protein matter, it is characterized in that its aminoacid sequence shown in SEQ ID NO:2, or this sequence is replaced, disappearance or add one or several amino-acid residue and the amino acid polypeptide with identical function that forms.

2, protein according to claim 1 is characterized in that aminoacid sequence is shown in SEQ ID NO:2.

3, the nucleotide sequence of the described proteinic resistance gene Pi 37 against rice blast of coding claim 1.

4, resistance gene Pi 37 against rice blast 6 according to claim 3, its nucleotide sequence is shown in SEQID NO:1.

5, a kind of carrier that contains claim 3 or 4 described genes.

6, the described carrier transgenic plant transformed of claim 5.

7, the molecule marker of claim 3 or 4 described genes generations.

8, claim 1 or the 2 described protein application in preparation resisting rice blast bacteria medicine.

9, the described molecule marker of claim 7 has application in the paddy rice of disease resistance in seed selection to rice blast.

Images