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Rice anti-rice-blast gene Pi36 co-dominant molecular marker and application

A rice blast resistance gene and molecular marker technology, applied in the direction of DNA/RNA fragments, microbial measurement/inspection, recombinant DNA technology, etc., can solve the problems of labor and cost, cumbersome resistance identification, inaccurate results, etc., to achieve The effect of low cost, shortened breeding cycle and simple operation

Active Publication Date: 2019-03-15
YUAN LONGPING HIGH TECH AGRI CO LTD +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In conventional disease resistance breeding, the identification of resistance is cumbersome, requires a lot of manpower and cost, and the identification is easily affected by external conditions, resulting in inaccurate results, so the breeding efficiency is low

Method used

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  • Rice anti-rice-blast gene Pi36 co-dominant molecular marker and application
  • Rice anti-rice-blast gene Pi36 co-dominant molecular marker and application
  • Rice anti-rice-blast gene Pi36 co-dominant molecular marker and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Example 1 Development of Pi36 gene molecular marker

[0032] The rice blast resistance gene Pi36 is located on the short arm of rice chromosome 8. By sequencing the genomic regions of Pi36 in rice varieties known to be resistant to or susceptible to rice blast, such as Kasalath, 9311, and Nipponbare, and online comparison analysis of the NCBI database, two specific genes were found. Sex SNP locus, wherein SNP1 is located at base 2886707 of chromosome 8 of Nipponbare sequence, Pi36 disease-resistant allele is G, and susceptible allele is A (such as figure 1 A); SNP2 is located at chromosome 2888118 of Nipponbare sequence, the resistant allele of Pi36 is G, and the susceptible allele is A (such as figure 1 shown in B). Two pairs of dominant markers were developed for these two SNP loci, which were used to identify the resistant allele and the susceptible allele respectively. The combined use of the two pairs of dominant markers could realize the identification of differe...

Embodiment 2

[0033] Example 2 Design of primers for detection of Pi36 gene co-dominant molecular markers

[0034] Detection primers were designed for the two Pi36 gene-specific molecular markers developed in Example 1, and the primer design principles were as follows.

[0035] First, the specific primers for amplifying the disease-resistant allele were designed, and the primer Pi36-RR was designed at the 3' end of the reverse primer with the complementary base C of the disease-resistant allele G at the SNP1 site, and at the 3' end of the primer Pi36-RR. At the 3rd base, the C base was changed to the A base to introduce a mismatch, and then the matching forward primer Pi36-RF was designed upstream of the base, and the primers Pi36-RR and Pi36-RF (SEQ ID NO: 3-4 ) can only specifically amplify the Pi36 disease-resistant allele, resulting in a 345bp band, while the Pi36 susceptible allele has no band during amplification;

[0036] Then, design specific primers to amplify the susceptible alle...

Embodiment 3

[0040] Example 3 Establishment of a specific molecular marker detection method for rice blast resistance gene Pi36

[0041] According to the detection primers of the two molecular markers of Pi36 designed in Example 2, the reaction program and reaction system of PCR were designed, and the following reaction program and system were determined through continuous optimization:

[0042] PCR reaction system (10μL): 1ul 10× PCR reaction buffer, 0.8ul 10mM dNTP, 4 primers (10μM) are 0.15μL, 0.1ul Taq DNA polymerase (2.5U / ul), 2ul DNA template, ddH 2 O supplemented to 10ul.

[0043] PCR reaction conditions were: pre-denaturation at 95°C for 5 minutes; denaturation at 95°C for 30 seconds, annealing at 56°C for 30 seconds, extension at 72°C for 45 seconds, a total of 32 cycles; extension at 72°C for 8 minutes.

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Abstract

The invention provides a rice anti-rice-blast gene Pi36 co-dominant molecular marker. The rice anti-rice-blast gene Pi36 co-dominance molecular marker comprises a marker SNP1 and / or a marker SNP2, wherein a polymorphic site of the marker SNP1 is located at the position of a basic group corresponding to a rice Nipponbare No. 8 chromosome 2886707, and the basic group at the position is G or A; a polymorphic site of the marker SNP2 is located at the position of a basic group corresponding to a rice Nipponbare No. 8 chromosome 2888118, and the basic group at the position is G or A. For the molecular marker, specific amplification primers are separately designed, and Pi36 genotype detection is carried out through PCR amplification. The molecular marker is co-dominant, three different genotypescan be distinguished effectively, the molecular marker can be used for breeding anti-rice-blast rice, the breeding period of the anti-rice-blast rice is shortened, the breeding speed is increased, andthe breeding cost is reduced. The rice anti-rice-blast gene Pi36 co-dominant molecular marker has the advantages of simplicity in operation, low cost, short period and the like.

Description

technical field [0001] The invention belongs to the field of molecular biology and plant molecular breeding, and in particular relates to a co-dominant molecular marker of rice blast resistance gene Pi36 and its application. Background technique [0002] Rice, as the main food and cash crop in my country, is threatened by diseases and insect pests all year round, among which rice blast is one of the most serious diseases. Rice blast occurs every year in various rice producing areas in China. When the disease and insect pests prevail, the yield can be reduced by 10% to 20%, and in more serious areas, it can reach 40% to 50%, or even no harvest. Extensive use of blast-resistant pesticides can alleviate the damage of rice blast, but it also greatly increases the cost of rice production and pollutes the environment. Practice has shown that breeding blast-resistant varieties is the most economical and effective measure to control rice blast. However, due to the large number of p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 杨远柱邓钊刘兰兰王凯秦鹏符辰建严天泽周延彪
Owner YUAN LONGPING HIGH TECH AGRI CO LTD
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