CN104404152B - Detect genotypic expression molecular labeling and the application of rice blast fungus basal resistance - Google Patents
Detect genotypic expression molecular labeling and the application of rice blast fungus basal resistance Download PDFInfo
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- CN104404152B CN104404152B CN201410723122.3A CN201410723122A CN104404152B CN 104404152 B CN104404152 B CN 104404152B CN 201410723122 A CN201410723122 A CN 201410723122A CN 104404152 B CN104404152 B CN 104404152B
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Abstract
The present invention provides the genotypic expression molecular labeling of detection rice blast fungus basal resistance and applications, can be used for identification of the rice to Pyricularia oryzae basal resistance.The nucleotides sequence of the molecular labeling is classified as SEQ ID No.1, using primer pair of the present invention, carries out PCR amplification by template of oryza sativa genomic dna, available and rice marks the genotype molecule of Pyricularia oryzae basal resistance gene close linkage.The presence or absence of the label, which can be specified, by PCR amplification analysis may determine that rice to the presence or absence of Pyricularia oryzae basal resistance, can be used in rice to Pyricularia oryzae basal resistance whether there is or not detection.The features such as genotype molecule marker bands are simple, distribution the is closeer advantage incomparable with SSR, quickly can accurately detect interracial polymorphism in subspecies, have broad application prospects.
Description
Technical field
The present invention relates to biomolecular labeling fields, in particular to detect the genotype table of rice blast fungus basal resistance
Up to molecular labeling and application.
Background technique
Genotype refers to the genotype of biology, that is, controls the assortment of genes type of character, be that organism is obtained from its parent
Obtain the summation of full gene.The character of one organism is very much, and the full gene for controlling these characters is known as biology
Body genotype.But the related gene composition that organism is studied character is generally referred to, it is the internal factor of trait expression, gene
Type is identified by hybrid experiment.If the genotype of the high stalk of pea can be indicated with DD or Dd, available dd of short stem is indicated.
It has been exploited for plant resistance to environment stress identification in spite of a large amount of molecular marking technique, but for passing through exploitation base
Because type molecular labeling carrys out the molecular labeling of plant identification resistance, there is presently no reports, since genotype determines phenotype, in addition I
Existing chip technology have found a large amount of genotype be it is related to rice basal resistance, therefore, exploitation genotype molecule marks
The accurate Rapid identification plant basal resistance of energy, more can direct accurate judgement rice basal resistance compared with other neutral molecules label.
Summary of the invention
To solve the above-mentioned problems of the prior art, the purpose of the present invention is to provide detection rice blast fungus bases
The genotypic expression molecular labeling of resistance and application can be used for identification of the rice to Pyricularia oryzae basal resistance.Utilize the present invention
Primer pair, using oryza sativa genomic dna as template carry out PCR amplification, available and rice is to Pyricularia oryzae basal resistance base
Because the genotype molecule of close linkage marks, which is named as M3 in the present invention.It can be clearly somebody's turn to do by PCR amplification analysis
The presence or absence of label may determine that rice to the presence or absence of Pyricularia oryzae basal resistance, can be used in rice to Pyricularia oryzae basal resistance
Whether there is or not detection.The features such as genotype molecule marker bands are simple, distribution the is closeer advantage incomparable with SSR, can be fast
Interracial polymorphism in fast accurate detection subspecies, has broad application prospects.It will be understood by those skilled in the art that in water
In the anti-pest identification of rice, is obtained outside molecular labeling of the invention except through the method for above-mentioned PCR amplification, tradition can also be passed through
The anti-pest identification methods of rice artificial infection rice varieties obtain molecular labeling of the invention.Traditional Rice Resistance pest identification
Method is easy to be influenced by environmental condition especially temperature, humidity and human factor, and molecular labeling of the invention can overcome
The influence of environmental condition improves the accuracy and efficiency of identification.
In order to achieve the above objectives, the technical solution of the present invention is as follows:
A kind of genotypic expression molecular labeling detecting rice blast fungus basal resistance, the nucleotides sequence of the molecular labeling
It is classified as SEQ ID No.1.
Further, the primer pair of the genotypic expression molecular labeling of the detection rice blast fungus basal resistance are as follows:
Prim3-F:TCGCATCAGCATTCAACGAAC；
Prim3-R:CGAAGAGGTTCTCGCCGTAG.
Further, the method for the genotypic expression molecular labeling of the detection rice blast fungus basal resistance are as follows: with
Disease-resistant and susceptible rice total dna is template, carries out PCR amplification with above-mentioned primer, PCR amplification condition be 95 DEG C of initial denaturation 3min,
94 DEG C of denaturation 1min, 55 DEG C of annealing 1min, 72 DEG C of extensions 1min, 72 DEG C of extension 10min.Amplified production is through 3% Ago-Gel
Electrophoresis detection compares the size of purpose band with molecular labeling, and the rice containing molecular labeling has to Pyricularia oryzae
Basal resistance.
Further, application of the molecular labeling in terms of the detection of rice blast fungus basal resistance and identification.
Compared with the existing technology, the invention has the benefit that
The present invention can be used for the identification of rice basal resistance with genotype molecule label of the rice to Pyricularia oryzae basal resistance.
The characteristics of according to rice to Pyricularia oryzae basal resistance candidate gene sequence, certain genotype molecule can be developed
Label.The method that the molecular labeling solves conventional method identification rice basal resistance avoids conventional method identification rice base
It is influenced when plinth resistance by human factor when environmental condition and inoculation and keeps qualification result inaccurate, molecular labeling of the invention
With it is quick, accurate, not by the interference of environment and human factor, can quickly understand whether identified rice varieties have basis
Resistance.The present invention can be used for the excavation etc. of resistant gene in plant with rice to the genotype molecule label of Pyricularia oryzae basal resistance
Aspect.Molecular labeling through the invention can sufficiently excavate rice basal resistance resource, to solve Rice Resistance pest breeding resources
Scarcity provides important information.
Detailed description of the invention
Fig. 1 is genotype molecule label detection rice varieties liyub and Lijiang xintuanheigu rice varieties electrophoretogram.
Wherein, M Marker, swimming lane 1 are rice varieties Liyub, and swimming lane 2 is rice varieties Lijiang xintuanheigu, swimming lane 3
For negative control.
Specific embodiment
With reference to the accompanying drawing and specific embodiment is described in further detail the present invention program:
Rice varieties used in the present invention are Yunnan tradition local varieties moon paddy, which is a middle water resistant rice product
Kind, it is inoculated with after the rice varieties by the strong pathogenic strain of Pyricularia oryzae and extracts rice total RNA, obtained Pyricularia oryzae and infect moon paddy
Transcript profile chip finds moon paddy when being infected 12h by Pyricularia oryzae, the expression quantity of gene is greatly by largely analyzing
Up-regulation.On this basis, genotypic expression molecular labeling M3 used in the present invention is devised, amplifying rice gene is used for
Os07g0124900.Specific step is as follows:
(1) genomic DNA and the susceptible rice varieties Lijing for extracting a disease resisting rice kind moon paddy respectively are new
The genomic DNA of the black paddy of group
(2) using genotypic expression molecular labeling Prim3 primer pair respectively to disease-resistant rice varieties genomic DNA and susceptible
Rice varieties genomic DNA carry out PCR amplification, PCR amplification condition be 95 DEG C of initial denaturation 3min, 94 DEG C of denaturation 1min, 55 DEG C move back
Fiery 1min, 72 DEG C of extensions 1min, 72 DEG C of extension 10min.
(3) with 3% Ago-Gel separation detection PCR product.
(4) pcr amplification product is sequenced, gained sequence is compared, and analyzes detected disease resisting rice kind DNA and expands
Whether whether there is or not the genotypic expression molecule marks involved in the present invention obtained for volume increase object genotype and susceptible rice varieties DNA cloning sequence
Note, to judge rice to the presence or absence of Pyricularia oryzae basal resistance.
As shown in Figure 1, for genotype molecule of the present invention label detection rice varieties liyub and Lijiang xintuanheigu rice product
Kind electrophoretogram.
Wherein, M Marker, swimming lane 1 are rice varieties Liyub, and swimming lane 2 is rice varieties Lijiang xintuanheigu, swimming lane 3
For negative control.
The above description is merely a specific embodiment, but scope of protection of the present invention is not limited thereto, any
The change or replacement expected without creative work, should be covered by the protection scope of the present invention.Therefore, of the invention
Protection scope should be determined by the scope of protection defined in the claims.
Claims (1)
1. a kind of genotypic expression molecular labeling for detecting rice blast fungus basal resistance, which is characterized in that the molecular labeling
Nucleotides sequence be classified as SEQ ID No.1；
The primer pair of the genotypic expression molecular labeling of the detection rice blast fungus basal resistance are as follows:
Prim3-F:TCGCATCAGCATTCAACGAAC；
Prim3-R:CGAAGAGGTTCTCGCCGTAG；
The method of the genotypic expression molecular labeling of the detection rice blast fungus basal resistance are as follows: with disease-resistant and susceptible rice
Total DNA is template, carries out PCR amplification with above-mentioned primer, PCR amplification condition is 95 DEG C of initial denaturation 3min, 94 DEG C of denaturation 1min, 55
DEG C annealing 1min, 72 DEG C of extensions 1min, 72 DEG C of extension 10min, amplified production is through 3% agarose gel electrophoresis detection PCR product；
Pcr amplification product is sequenced, gained sequence is compared, and analyzes detected disease resisting rice kind DNA cloning product gene
Whether whether there is or not the involved genotypic expression molecular labelings obtained for type and susceptible rice varieties DNA cloning sequence, to judge rice pair
The presence or absence of Pyricularia oryzae basal resistance.
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| CN105755157A (en) * | 2016-05-11 | 2016-07-13 | 东北农业大学 | PCR (Polymerase Chain Reaction) detection primer group for magnaporthe grisea and detection method thereof |
| CN110257473A (en) * | 2019-07-01 | 2019-09-20 | 湖北省农业科学院植保土肥研究所 | Resistant rice varieties plant blast resisting identification method in specific rice region |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1840696A (en) * | 2006-01-26 | 2006-10-04 | 中国水稻研究所 | Molecule identification and transfer technology for broad-spectrum rice-blast resistant gene of paddy rice |
| CN101906427A (en) * | 2010-06-25 | 2010-12-08 | 华南农业大学 | Rice blast resistance gene Pi1 and application thereof |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1840696A (en) * | 2006-01-26 | 2006-10-04 | 中国水稻研究所 | Molecule identification and transfer technology for broad-spectrum rice-blast resistant gene of paddy rice |
| CN101906427A (en) * | 2010-06-25 | 2010-12-08 | 华南农业大学 | Rice blast resistance gene Pi1 and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| 4 个抗稻瘟病基因分子标记的建立及在水稻亲本中的分布;高利军;《第1届中国杂交水稻大会论文集》;20100930;294-298 |
| Development of Dominant Rice Blast Pi-ta Resistance Gene Markers;Yulin Jia;《Crop Science》;20021231;第42卷;2145-2149 |
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