CN106048069A - Molecular markers for detection of rice broad-spectrum rice blast-resistant gene Pike and application thereof - Google Patents
Molecular markers for detection of rice broad-spectrum rice blast-resistant gene Pike and application thereof Download PDFInfo
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Abstract
The invention discloses molecular markers for detection of a rice broad-spectrum rice blast-resistant gene Pike and an application thereof, and belongs to the field of molecular biology. The molecular markers disclosed by the invention comprise a molecular marker CP-G1328C containing DNA fragments 1, 2, 3, 4 and 6, a molecular marker CP-G1328T containing DNA fragments 1, 2, 4, 5 and 6, and a molecular marker CP-G1328G' containing DNA fragments 1, 2, 4, 6 and 7, wherein the DNA fragments 1-7 have the sequences shown in SEQ ID NO.1-7 respectively. Primers for amplifying the molecular markers comprise Pik-F, Pik-R, G-F, C-R, T-R and G'-R, and have the sequences respectively shown in SEQ ID NO.8-13. The molecular markers or primers can be used for identification of the rice broad-spectrum rice blast-resistant gene Pike allele type, screening of germplasm resources and molecular marker assisted selection breeding.
Description
Technical field
The invention belongs to biology field, relate to detect broad-spectrum rice-blast resistant gene of paddy rice Pike molecular marker and
Its application.
Background technology
Rice blast (Magnaporthe Oryzae) is one of important disease of Oryza sativa L. (Oryza sativa L.), full generation
Boundary there are about the rice yield of 10%-30% every year and suffers a loss due to the invasion of Pyricularia oryzae.Production practices show, utilize anti-
It is rice blast method most economical, effective of preventing and treating at present that rice blast gene cultivates paddy disease-resistant new varieties.Along with to Rice Resistance rice
The research of pestilence molecular mechanism, has had more than 20 blast resistant gene to be cloned, and wherein Pike is positioned at Oryza sativa L. o.11
Chromosome long arm end, is the neomorph on resistance locus Pik, and this gene is from long-grained nonglutinous rice strain Hunan 143 (Hunan Province Oryza sativa L. morning
Institute) middle clone, inoculated identification finds that this gene pairs rice blast has broad spectrum durable resistance, by MAS (molecular marker
Assisted Selection) etc. means will this channel genes to R287, Japan's high-grade rice strain such as fine find, this gene can be greatly improved
The rice blast resistance of rice strain.So, Pike has the highest using value in the breeding for disease resistance of Oryza sativa L..
In rice genome, the rice blast resistance of disease-resistant gene Pike is by two adjacent NBS-LRR genoid Pike-
1 and Pike-2 together decides on, and utilizes Multiple Sequence Alignment software DNAMAN8.0 by Pike coding region sequence and other 6 Pik equipotentials
Gene (Pik, Pikh, Pikm, Pikp, Piks, Pi1) compares, and finds to also exist the 1328th of Pike-1cDNA sequence
One specific SNP site G1328C of Pike.The nucleotide that this SNP site is corresponding in Pike is G, and corresponding aminoacid is
Tryptophan (W), and this SNP site in other 6 allele cloned (Pik, Pikh, Pikm, Pikp, Piks, Pi1)
Corresponding nucleotide is C, and coded amino acid is serine (S), so the ucleotides corresponding by detecting SNP site G1328C
Type can effectively distinguish Pike and other 6 the Pik allele cloned.But, find SNP site in nature by research
Nucleotide type corresponding for G1328C not only exists G, C two types, and existing molecular marker can not be to SNP site G1328C
The type of other corresponding nucleotide effectively detects.
Summary of the invention
The present invention has randomly selected the hybrid paddy rice backbone parent materials such as Japanese fine, Yue Tai B, awns be extensive and to Pike-1 equipotential base
The partial nucleotide sequence of cause has carried out sequencing analysis, finds the ucleotides corresponding according to SNP site C1328C in Pike-1
The allelic variation of Pike in nature can be divided into 4 type (G-type, C-type, T-by type and flanking region sequence thereof
Type, G '-type), and in sequence alignment discovery same kind, Pike allelic nucleotide sequence similarity is higher, and not
Relatively low with the nucleotide sequence similarity between type.The present invention is according to this sequencing results simultaneously, utilizes SNP site
Nucleotide that G1328C is corresponding and SNP site flanking nucleotide sequences have developed the codominance of 3 specific detection Pike
Molecular marker, these codominant markers have specificity to the detection of blast resistant gene Pike, can effectively distinguish simultaneously
The allelic variation of Pike site 4 type.Compared with the dCAPS labelling d-G1328C of specific detection Pike of forefathers' exploitation,
This codominant marker has that testing cost is cheap, simple to operate, the feature of applied range.Simultaneously as these 3 molecular markers
Being positioned at the inside of Pike, in MAS breeding, these molecular markers and disease-resistant gene Pike are divided into from, and Pike linksystem molecule
The testing result of labelling RM224 is compared, and these labellings are the most accurate to the testing result of Pike, reliable.
It is an object of the invention to provide the molecular marker of 3 detection broad-spectrum rice-blast resistant gene of paddy rice Pike.The present invention
Purpose also reside in the primer expanding described molecular marker be provided, and described molecular marker or the application of primer.
The purpose of the present invention is achieved through the following technical solutions:
The present invention has randomly selected 12 parts of hybrid rice breeding backbone parents including 93-11, Yue Tai B, and to these
Pike-1 allele coding region 1168bp-1828bp in material uses primer combination Pik-F and TG-R (table 1) to carry out
RCR expands, clones, checks order.Expand through PCR, from Peiai 64S, R287, Basmatis, good 814 and Lijiang xintuanheigu 5 parts
Obtaining the DNA fragmentation of a length of 661bp in material, sequence alignment finds, these 5 parts order-checking materials and Pik-1, Pikh-1,
Pikm-1, Pikp-1, Piks-1 and Pi1-5C sequence similarity in this region Yu Pike-1 has reached more than 99%, but
At SNP site G1328C, these 5 parts of materials and the nucleotide in Pikm-1, Piks-1, Pi1-1, Pik-1, Pikp-1, Pikh-1
Unanimously, all bases C (Fig. 1)；From 93-11, Yue Tai B, IR6, Huang Huazhan and closing, rich viscous 5 parts of materials obtain a length of
The DNA fragmentation of 640bp, sequence alignment finds, these 5 parts of materials and the Pike-1 sequence similarity in this region are only 77.6%-
78%, but reached 99% with the similarity of Pik-Nip (Japan fine middle Pik allele), at SNP site G1328C, these are 5 years old
Part material is consistent with the nucleotide in Pik-Nip, all bases T；A length of 670bp is obtained from Fuhui 838 and awns are extensive
DNA fragmentation, these 2 parts of materials and the Pike-1 sequence similarity in this region are only 73.5%, with the similarity of Pik-Nip are
73.8%, although at G1328C, come from the allele identical with Pike-1 (Fig. 1) of these 2 parts of materials, but in SNP position
Near point, its nucleotide sequence is relatively low with Pike similarity.
Convenient for subsequent descriptions, the allelic variation in Pik site is divided into 4 kinds according to above-mentioned the sequencing results by the present invention
Type: 1) base corresponding for G-type:SNP site G1328C is G, such as: Pike；2) C-type:SNP site G1328C is corresponding
Base is C, as: Pikm-1, Piks-1, Pi1-1, Pik-1, Pikp-1, Pikh-1 and derive from 5 parts of materials such as Peiai 64S
Pik allele；3) base corresponding for T-type:SNP site G1328C is T, such as Pik-Nip and derive from 5 parts of materials such as 93-11
The Pik allele of material；4) G '-type: although base corresponding to SNP site G1328C is G, but nucleotide sequence and Pike
Similarity relatively low, such as the Pik allele (Fig. 1) deriving from that awns is extensive and in Fuhui 838.
The present invention, according to nucleotide type corresponding to above-mentioned the sequencing results and SNP site G1328C, develops 3 energy
Codominant marker CP-G1328C, CP-G1328T, CP-of enough specific detection rice blast resistant gene Pike
G1328G’.Wherein, molecular marker CP-G1328C is to use primer combination Pik-F, Pik-R, G-F and C-R from rice genome
Middle amplification DNA fragmentation out, it comprises DNA fragmentation 1, DNA fragmentation 2, DNA fragmentation 3, DNA fragmentation 4 and DNA fragmentation 6, should
Molecular marker shows as codominance between Pike (G-type) and C-type allele；Molecular marker CP-G1328T is to use
Primer combination Pik-F, Pik-R, G-F and T-R DNA fragmentation of expanding out from rice genome, it comprises DNA fragmentation 1,
DNA fragmentation 2, DNA fragmentation 4, DNA fragmentation 5 and DNA fragmentation 6, this molecular marker is at Pike (G-type) and T-type allele
Between show as codominance；Molecular marker CP-G1328G ' is that use primer combination Pik-F, Pik-R, G-F and G '-R is from Oryza sativa L. base
Because of the DNA fragmentation expanding out in group, it comprises DNA fragmentation 1, DNA fragmentation 2, DNA fragmentation 4, DNA fragmentation 6 and DNA fragmentation
7, this molecular marker shows as codominance between Pike (G-type) and G '-type allele.The sequence of DNA fragmentation 1-7 is divided
Not as shown in SEQ ID NO.1-7.
Expand the primer sequence that the fragment that above-mentioned codominant marker comprises used as follows:
Pik-F:5 '-GAAGAATGGGAAGTCATCAGA-3 '；
Pik-R:5 '-ACCTTCTGCTGCTTTCTCTTC-3 '；
G-F:5 '-GATCTAGATAATAATGATGCTTTGTG-3 '；
C-R:5 '-GCTATCCTCCAAGACATCGA-3 '；
T-R:5 '-CTTGCTACCTTGTCACACAACA-3 '；
G '-R:5 '-AGACCTCTCTATTGTACTGT-3 '.
Above-mentioned codominant marker or primer differentiate at rice blast resistant gene Pike allelic gene type, germ plasm resource sieve
Application in choosing and molecular marker assisted selection breeding.
For codominant marker CP-G1328C, labelling CP-G1328T and labelling CP-G1328G ', no matter which kind of class
The allele of type exists, and two external primers Pik-F and Pik-R can amplify a total fragment, wherein G-type and
C-type allelic total fragment a length of 558bp, T-type and G ' the allelic total fragment length of-type is respectively
549bp and 576bp.For molecular marker CP-G1328C, in the presence of G-type allele, primer combination G-F and Pik-R
Can amplify G-type allele-specific fragment, this fragment length is 423bp；And in the presence of C-type allele,
Primer combination Pik-F and C-R can amplify the peculiar fragment of C-type allele of an a length of 179bp.Should although using
During 4 kinds of allele of marker detection, the DNA fragmentation length of amplification there are differences, but is using low concentration agarose gel to expansion
When volume increase thing detects, this molecular marker can only show altogether between the two kinds of allele of G-type and C-type
Dominant.
Next to that codominant marker CP-G1328T, in the presence of G-type allele, primer combination G-F and
Pik-R can expand the G-type allele-specific fragment of a length of 423bp of shaping, does not then have for C-type allele
Specific fragment is had to be amplified out.For T-type allele, primer combination Pik-F and T-R can amplify one
The T-type allele-specific fragment of 182bp.In actual applications, molecular marker CP-G1328T only can at G-type and
Codominance is shown between T-type allele.
For the 3rd codominant marker CP-G1328G ', this is marked at institute in G-type and C-type allele
DNA fragmentation and the molecular marker CP-G1328T that can amplify are completely the same.But in the presence of working as G '-type allele, primer
Combination G '-R and Pik-F can amplify an allele specific DNA fragmentation of G '-type, and this DNA fragmentation is a length of
192bp.So, in actual applications, molecular marker CP-G1328G ' is only capable of at Pike (G-type) and G '-type allele
Between show codominance, and show dominant between other combine.
Although above-mentioned 3 molecular markers are based on same SNP site and flanking region developed, can effectively examine
Survey Pike (G-type), but its performance between Pike difference allelic gene type is different, so needing root in actual applications
Select according to the allelic type of Pike contained in rice material.
The present invention has such advantages as relative to prior art and effect: the codominant marker primer energy of the present invention
Enough the most specific oryza sativa genomic dna is marked qualification, wide spectrum blast resistant gene in detection rice material genomic DNA
The existence (isozygotying or heterozygosis) of Pike, detection accuracy is high, with the method phase such as traditional enzyme action, order-checking and anti-disease enzyme
Ratio, have with low cost, time-consuming less, advantage simple to operate.
Accompanying drawing explanation
Nucleotide sequence comparison result figure near Fig. 1 SNP site G1328C.Pik-1、Pikh-1、Pikm-1、Pikp-1、
Piks-1, Pi1-5C are the allele having cloned 6 Pike-1.SNP site G1328C is as shown in black box.
Fig. 2 detects the molecular markers development schematic diagram of Pike allelic gene type.
Fig. 3 detects the agarose gel electrophoresis figure of codominant marker design result.A: codominant marker CP-
The testing result of G1328C；B: the testing result of codominant marker CP-G1328T；C: codominant marker CP-
The testing result of G1328G '；The testing result of d:dCAPS labelling d-G1328C.M:DL2000 (TaKaRa, Janpan)；1: Hunan
Early 143 (Pike)；2:IRBLkm (Pik-m)；3:IRBLk-ka (Pik)；4:IRBLks-F5(Pik-s)；5:IRBL1-CL
(Pi1)；6:IRBLkp-K60 (Pik-p)；7:IRBLkh-K3 (Pik-h)；8: Lijiang xintuanheigu；9: Japan is fine；10: Peiai
64S；11: Fuhui 838；12: Yue Tai B；13:IR6；14:Basmatis；15: good 814；16: close rich viscous；17: awns is extensive；18:
R287。
The electrophoretogram of Fig. 4 Markers for Detection rice breeding strain genotype.A molecular marker RM224 is to 14 parts of breeding strains
It it is the electrophoretogram of genotype detection；The b molecular marker CP-G1328C agarose gel electricity to 14 parts of breeding strain genotype detection
Swimming figure.In figure from left to right, M: molecular weight marker DL2000 (TaKaRa, Japan)；1: Hunan early 143 (Pike donors)；2:
R287；3:75-1-127；4:B505；5-18:HD1-HD14.
Detailed description of the invention
Below in conjunction with embodiment and accompanying drawing, the present invention is described in further detail, but embodiments of the present invention do not limit
In this.If not specializing, the conventional means that technological means used in embodiment is well known to those skilled in the art.
The present invention utilizes broad-spectrum rice-blast resistant gene of paddy rice Pike, Pi1, Pik, Piks, Pikm, Pikp, Pikh and part
In material, Pike-1 homologous sequence is at the nucleotide sequence difference (Fig. 1) of 1328, coding region and flanking region thereof, for SNP site
G1328C designs the molecular marker of 3 specific detection Pike (G-type), and each molecular marker contains 5 DNA fragmentations,
Wherein molecular marker CP1-G1328C contains DNA fragmentation 1, DNA fragmentation 2, DNA fragmentation 3, DNA fragmentation 4 and DNA fragmentation 6；Point
Sub-labelling CP-G1328T contains DNA fragmentation 1, DNA fragmentation 2, DNA fragmentation 4, DNA fragmentation 5 and DNA fragmentation 6；Molecular marker
CP-G1328G ' contains DNA fragmentation 1, DNA fragmentation 2, DNA fragmentation 4, DNA fragmentation 6 and DNA fragmentation 7.Relevant sequence dna fragment
As follows:
The sequence (the total DNA fragmentation of G-type and C-type, length 558bp, SEQ ID NO.1) of DNA fragmentation 1,
The sequence (Pike (G-type) peculiar fragment, length 423bp, SEQ ID NO.2) of DNA fragmentation 2,
The sequence (the peculiar fragment of C-type, length 179bp, SEQ ID NO.3) of DNA fragmentation 3,
The sequence (the peculiar fragment of T-type, length 549bp, SEQ ID NO.4) of DNA fragmentation 4,
The sequence (the peculiar fragment of T-type, length 182bp, SEQ ID NO.5) of DNA fragmentation 5,
The sequence of DNA fragmentation 6 (G ' the peculiar fragment of-type, length 576bp, SEQ ID NO.6),
The sequence of DNA fragmentation 7 (G ' the peculiar fragment of-type, length 192bp, SEQ ID NO.7).
According to the marker development principle shown in Fig. 2, according to 1328, coding region nucleotide type and SNP site flanking region
Sequence difference, devises the primer (table 1) of the molecular marker of specific detection Pike.
Primer sequence used in table 1 present invention and function information
The application in distinguishing Pike and other types allele of embodiment 1 codominant marker
The present invention uses donor material Hunan of Pike early 143, containing allelic 6 Monogenic lines: the IRBLkm-of Pike
Ts(Pikm)、IRBLk-Ka(Pik)、IRBLks-F5(Piks)、IRBL1-CL(Pi1)、IRBLkp-K60(Pikp)、IRBLkh-
K3 (Pikh), 10 target zone have checked order rice strain (Fig. 1): Lijiang xintuanheigu, Peiai 64S, Fuhui 838, Yue Tai B,
IR6, Basmatis, good 814, close rich viscous, awns is extensive, R287 and Japan fine (T-type), to the molecular markers development in the present invention
Result is verified, uses dCAPS labelling d-G1328C as comparison simultaneously.
Utilize CTAB method to extract the genomic DNA of above-mentioned rice material, DNA fragmentation contained by each molecular marker will be expanded
Article 4, primer (CP-G1328C:Pik-F, Pik-R, G-F, C-R；CP-G1328T:Pik-F, Pik-R, G-F, T-R；CP-
G1328G ': Pik-F, Pik-R, G-F, G '-R) respectively mixed in equal amounts, then by the primer after mixing as in PCR amplification system
Primer, the genomic DNA of above-mentioned material is carried out PCR amplification, PCR expands use 2 × ES Taq Master Mix
(CWBIO, Beijing, China), amplification condition is: after 94 DEG C of 5min denaturations, and 94 DEG C of 45s, 50 DEG C of 45s and 72 DEG C of 45s are even
Continuous 30 circulations；72 DEG C of 10min subsequently, 4 DEG C of 5min.PCR primer uses the agarose gel of 2% to detect, testing result
See Fig. 3.
From figure 3, it can be seen that for codominant marker CP-G1328C, labelling CP-G1328T and labelling CP-
G1328G ', the most what type of allele exists, and all can amplify a DNA fragmentation, and wherein G-type (compile by swimming lane
Number 1) and C-type (swimming lane numbering 2-8,10,14,15,18) a length of 558bp of allelic DNA fragmentation, T-type (swimming lane is compiled
Numbers 9,12,13,16) and G '-type (swimming lane numbering 11,17) allelic total fragment length be respectively 549bp and
576bp。
For codominant marker CP-G1328C, in the material Hunan containing allele Pike (G-type) early 143, should
Codominant marker can expand 1 the peculiar fragment of Pike (G-type) (423bp)；Containing the allelic material of C-type
In, it is possible to expand 1 the peculiar fragment of C-type allele (179bp) (Fig. 3 a).So this be marked at Pike (G-type) and
Codominance is shown as between C-type allele.
For codominant marker CP-G1328T, in the material Hunan containing allele Pike (G-type) early 143, should
Codominant marker can expand 1 the peculiar fragment of Pike (G-type) (423bp)；Containing the allelic material of T-type
In, it is possible to expand 1 the peculiar fragment of T-type allele (182bp) (Fig. 3 b).So this be marked at Pike (G-type) and
Codominance is shown as between T-type allele.
For codominant marker CP-G1328G ', in the material Hunan containing allele Pike (G-type) early 143, should
Codominant marker can expand 1 the peculiar fragment of Pike (G-type) (423bp)；Containing the allelic material of G '-type
In, it is possible to expand 1 the peculiar fragment of G '-type allele (192bp) (Fig. 3 c).So this be marked at Pike (G-type) and
Codominance is shown as between G '-type allele.
The above results shows, these codominant markers newly developed except can to Pike and other cloned 6
Individual allele (Pi1, Pik, Pikm, Piks, Pikp, Pikh) is effectively distinguished, moreover it is possible to effectively differentiation 4 kinds is different types of
Pike-1 allelic variation, compared with dCAPS labelling d-G1328C (Fig. 3 d), the range of application of these 3 molecular markers is wider and grasps
Make the simplest.
The application in Screening of Germplasm of embodiment 2 codominant marker
Another function of Middle molecule labelling of the present invention is the screening antigen-like material containing Pike from Rice Resources, for checking
This function, the present invention has randomly selected 320 parts of rice strains, to molecular marker CP-G1328C, CP-G1328T in the present invention
With the function of CP-G1328G ' verified.By Markers for Detection, in 320 parts of materials, find altogether containing G-type
Allelic material 1 part, containing the allelic material of C-type 129 parts, containing the allelic material of T-type 131
Part, containing the allelic material of G '-type 59 parts.
Owing to enzyme action is one of most effectual way of detection SNP site, dCAPS labelling d-G1328C is used to check this
The accuracy that 320 parts of material SNP site G1328C are detected by the molecular marker in bright, result shows the molecular marker pair of the present invention
The testing result of above-mentioned material is correct.Owing to d-G1328C can not in containing the allelic material of T-type and G '-type
Effectively amplification, the present invention has randomly selected 13 parts of T-type and 6 parts of G '-type rice strains are checked order, and sequencing result is demonstrate,proved
Real Middle molecule labelling of the present invention is correct to the testing result of above-mentioned material.
Owing to Middle molecule of the present invention is labeled as the codominant marker of specific detection Pike, for verification mark pair
The specificity of Pike detection, the present invention is 3 right to using containing the Pike-1 in G-type allele material 898B of filtering out
Amplified fragments has the primer of overlap and (KE-1F, KE-1R, KE-2F, KE-2R, KE-3F, KE-3R) (table 1) has been carried out PCR expansion
Increase, check order, and the nucleotide sequence of sequencing result with Pike is compared, it was found that rice strain 898B contains really
There is wide spectrum blast resistant gene Pike.
The above results shows, the molecular marker in the present invention can specific detection Pike, simultaneously dividing in the present invention
Sub-labelling can effectively distinguish the allele of 4 kinds of Pik site dissimilar (G-type, C-type, T-type, G '-type),
And amplified band clearly becomes clear, detection method is simple, so can effectively apply in the breeding for disease resistance of Oryza sativa L..
The application in molecular marker assisted selection breeding of embodiment 3 codominant marker
The present invention have chosen by Hunan 14 parts of rice breeding strains HD1-HD14 that early 143 (Pike antigens) are derivative, to this
The molecular marker in bright function in breeding for disease resistance is verified.First by molecular marker closely linked with Pike
Breeding strain is detected by RM224, and testing result shows all to contain Pike (Fig. 4 a) in 14 breeding strains.But use this
Molecular marker CP-G1328C in bright identifies and finds, without wide spectrum blast resisting base in breeding strain HD5, HD12, HD13, HD14
Because of Pike (Fig. 4 b).In order to verify the testing result of two labellings, use spontaneous induction method to R287, Hunan early in enshi
143 and breeding strain HD1-HD14 carried out rice blast resistance assessment (table 2), contrast Markers for Detection it was found that this
The genotype results that bright Middle molecule labelling is detected is consistent with Resistance Identification phenotypic results.This result shows, relative to Pike
The molecular marker RM224 of linksystem, the molecular marker in the present invention is the most accurate to the qualification result of Pike.
The assessment of table 2 rice breeding strain rice blast resistance and Markers for Detection result
R: disease-resistant；MR: in anti-；MS: middle sense；S: susceptible."+" Pike detects the positive；"-" Pike detection feminine gender.
Above-described embodiment is the present invention preferably embodiment, but embodiments of the present invention are not by above-described embodiment
Limit, the change made under other any spirit without departing from the present invention and principle, modify, substitute, combine, simplify,
All should be the substitute mode of equivalence, within being included in protection scope of the present invention.
Claims (3)
1. detection broad-spectrum rice-blast resistant gene of paddy ricePikeMolecular marker, it is characterised in that: for CP-G1328C, CP-
G1328T、CP-G1328G’；Wherein, molecular marker CP-G1328C contains DNA fragmentation 1, DNA fragmentation 2, DNA fragmentation 3, DNA
Fragment 4, DNA fragmentation 6；Molecular marker CP-G1328T contain DNA fragmentation 1, DNA fragmentation 2, DNA fragmentation 4, DNA fragmentation 5,
DNA fragmentation 6；Molecular marker CP-G1328G ' contains DNA fragmentation 1, DNA fragmentation 2, DNA fragmentation 4, DNA fragmentation 6, DNA fragmentation
7；The nucleotide sequence of DNA fragmentation 1-7 is respectively as shown in SEQ ID NO.1-7.
2. the primer of amplification molecular marker described in claim 1, it is characterised in that: for Pik-F, Pik-R, G-F, C-R, T-R,
G '-R, each primer sequence is as follows:
Pik-F:5 '-GAAGAATGGGAAGTCATCAGA-3 '；
Pik-R:5 '-ACCTTCTGCTGCTTTCTCTTC-3 '；
G-F:5 '-GATCTAGATAATAATGATGCTTTGTG-3 '；
C-R:5 '-GCTATCCTCCAAGACATCGA-3 '；
T-R:5 '-CTTGCTACCTTGTCACACAACA-3 '；
G '-R:5 '-AGACCTCTCTATTGTACTGT-3 '；
Wherein, primer combination Pik-F, Pik-R, G-F and C-R for amplifier molecule labelling CP-G1328C, primer combination Pik-F,
Pik-R, G-F and T-R are used for amplifier molecule labelling CP-G1328T, primer combination Pik-F, Pik-R, G-F and G '-R is used for expanding
Molecular marker CP-G1328G '.
3. the molecular marker described in claim 1 or the primer described in claim 2 are at rice blast resistant genePikeEquipotential
Application in gene type discriminating, Screening of Germplasm, molecular marker assisted selection breeding.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107904324A (en) * | 2017-12-30 | 2018-04-13 | 武汉大学 | The codominant marker system of avirulence gene of rice blast AVR Pik and its primer and application |
| CN111978387A (en) * | 2020-08-26 | 2020-11-24 | 武汉大学 | Rice blast resistance gene Pikg, encoding protein and application thereof |
| CN113999934A (en) * | 2021-12-14 | 2022-02-01 | 湖北省农业科学院粮食作物研究所 | Rice blast resistance Pik locus allele identification molecular marker and application thereof |
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| CN104531717A (en) * | 2014-12-18 | 2015-04-22 | 武汉大学 | Rice-blast resistant genes Pike and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104531717A (en) * | 2014-12-18 | 2015-04-22 | 武汉大学 | Rice-blast resistant genes Pike and application thereof |
Non-Patent Citations (1)
| Title |
|---|
| CHEN ET AL: "Pike, a rice blast resistance allele consisting of two adjacent NBS-LRR genes, was identified as a novel allele at the Pik locus.", 《MOLECULAR BREEDING》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107904324A (en) * | 2017-12-30 | 2018-04-13 | 武汉大学 | The codominant marker system of avirulence gene of rice blast AVR Pik and its primer and application |
| CN111978387A (en) * | 2020-08-26 | 2020-11-24 | 武汉大学 | Rice blast resistance gene Pikg, encoding protein and application thereof |
| CN111978387B (en) * | 2020-08-26 | 2021-11-02 | 武汉大学 | Rice blast resistance gene Pikg, encoding protein and application thereof |
| CN113999934A (en) * | 2021-12-14 | 2022-02-01 | 湖北省农业科学院粮食作物研究所 | Rice blast resistance Pik locus allele identification molecular marker and application thereof |
| CN113999934B (en) * | 2021-12-14 | 2022-08-05 | 湖北省农业科学院粮食作物研究所 | Rice blast resistance Pik locus allele identification molecular marker and application thereof |
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