General codominance molecular marker for resisting nilaparvata lugens Stal BPH9 multiple alleles of rice, and detection method and application of general codominance molecular marker

An anti-BPH and allele technology, applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., can solve the problems of loss of resistance, etc., and achieve efficient detection, efficient and accurate screening, and detection procedures Convenient and fast effect

Active Publication Date: 2019-04-02
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

However, due to the migratory nature of N. lugens and different biotypes, rice varieties contain

Method used

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  • General codominance molecular marker for resisting nilaparvata lugens Stal BPH9 multiple alleles of rice, and detection method and application of general codominance molecular marker
  • General codominance molecular marker for resisting nilaparvata lugens Stal BPH9 multiple alleles of rice, and detection method and application of general codominance molecular marker
  • General codominance molecular marker for resisting nilaparvata lugens Stal BPH9 multiple alleles of rice, and detection method and application of general codominance molecular marker

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1 Development of rice universal co-dominant molecular markers for resistance to brown planthopper multiple alleles BPH1 / 10 / 18 / 21, BPH2 / 26, BPH7 and BPH9

[0043] Rice BPH-resistant multiple alleles BPH1 / 10 / 18 / 21, BPH2 / 26, BPH7, and BPH9 are located on the long arm of rice chromosome 12, and can resist BPH parents Mudgo (BPH1 / 10 / 18 / 21), ADR52 (BPH2 / 26), T12 (BPH7), Pokkali (BPH9), and the susceptible parents 9311, Nipponbare, Huazhan, 638S and more than 20 kinds of rice material genome sequence determination and comparison analysis, in the third exon of the brown planthopper resistance gene A specific MNP (polynucleotide polymorphism) was found, located at 22874098-22874099 bp on chromosome 12 of Nipponbare, in which BPH1 / 10 / 18 / 21, BPH2 / 26, BPH7, BPH9 insect resistance alleles were 5'- TG-3', the susceptible allele is 5'-CC-3' or 5'-CA-3'. The sequence analysis results of the four types of BPH-resistant multiple alleles BPH7, BPH9, BPH18, BPH26 and 9311, Nippon...

Embodiment 2

[0044] Embodiment 2 Multiple alleles BPH1 / 10 / 18 / 21, BPH2 / 26, BPH7, design of detection primers for co-dominant molecular markers of BPH9 genes

[0045] Detection primers were designed for the multiple allele-specific molecular markers developed in Example 1, and the principle of primer design was as follows.

[0046] Firstly, specific primers for amplifying the brown planthopper-resistant allele (hereinafter referred to as the insect-resistant allele) were designed, and the 5 alleles of the insect-resistant multiple alleles BPH1 / 10 / 18 / 21, BPH2 / 26, BPH7, and BPH9 were designed. The 5'-CA-3' complementary to '-TG-3'MNP is used to design the reverse primer BPH1 / 9-RR at the 3' end of the primer, and then design the forward primer BPH1 / 9-RF paired with it upstream, Primers BPH1 / 9-RR and BPH1 / 9-RF can only specifically amplify BPH1 / 10 / 18 / 21 or BPH2 / 26 or BPH7 or BPH9 to produce 360bp bands, but no amplification bands for other alleles;

[0047] Then, design specific primers for amp...

Embodiment 3

[0051] Example 3 Establishment of a universal molecular marker detection method for rice resistance to brown planthopper multiple alleles BPH1 / 10 / 18 / 21, BPH2 / 26, BPH7 and BPH9

[0052] According to the detection primers of BPH1 / 10 / 18 / 21, BPH2 / 26, BPH7 and BPH9 universal molecular markers designed in Example 2, the reaction program and reaction system of PCR were designed, and through continuous optimization, the following reaction program and system were determined:

[0053] PCR reaction system (10 μL): 1 μL of 10×PCR reaction buffer, 0.8 μL of 10 mM dNTP, 0.15 μL of each of the 4 primers (10 μM), 0.1 μL of Taq DNA polymerase; 2 μL of DNA template, and double distilled water to make up the rest.

[0054] The 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 35 cycles; extension at 72°C for 8 minutes.

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Abstract

The invention relates to a general codominance molecular marker for resisting nilaparvata lugens Stal BPH9 multiple alleles of rice, and a detection method and application of the general codominance molecular marker. The general molecular marker for resisting nilaparvata lugens Stal multiple alleles BPH1/10/18/21, BPH2/26, BPH7 and BPH9 of the rice, provided by the invention, is located at (22874098bp)th-(22874099bp)th position of a 12th chromosome of the rice, and the polymorphism is 5'-TG-3'/5'-CC-3'/5'-CA-3'. The general codominance molecular marker for resisting nilaparvata lugens Stal multiple alleles of rice provided by the invention can be generally used for identification of homozygous genotypes or heterozygous genotypes of the multiple alleles BPH1/10/18/21, BPH2/26, BPH7 and BPH9, and efficient accurate identification of BPH1/10/18/21, BPH2/26, BPH7 and BPH9 genotypes of nilaparvata lugens Stal resisting rice recourse is realized.

Description

technical field [0001] The invention relates to the field of plant molecular breeding, in particular to the universal co-dominant molecular markers of multiple alleles BPH1 / 10 / 18 / 21, BPH2 / 26, BPH7, BPH9 in rice resistant to brown planthopper BPH9 and its detection method and application. Background technique [0002] Brown planthopper (Nilaparvata lugens Stal, referred to as BPH) belongs to Homoptera planthopper insects, is the main pest of rice. Brown planthoppers are widely distributed in my country, and have strong seasonality, migration and reproductive ability. Outbreaks often cause rice plants to wither, yield reduction or even failure. Rice planthoppers also spread rice diseases while feeding on rice, aggravating the damage to rice and causing greater losses to rice planting. Existing studies have shown that cultivating brown planthopper-resistant rice varieties using rice's own resistance is one of the most effective, economical and safest measures to control brown ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 杨远柱邓钊何光存郭建平王凯严天泽秦鹏符辰建
Owner YUAN LONGPING HIGH TECH AGRI CO LTD
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