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Molecular marker detection method of rice blast bacterium non-toxic genes Avr-Pit and primers thereof

The technology of avr-pit and rice blast fungus is applied in the field of molecular markers for detecting the avirulent gene Avr-Pit of rice blast fungus, which can solve the problems of complex operation procedures, biochemical reagents, inconvenient and avirulent genotoxicity composition analysis, etc., and achieve clear results. , low accuracy, saving time and cost

Inactive Publication Date: 2012-09-05
NANJING AGRICULTURAL UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Most of the molecular markers linked to non-toxic genes (especially the molecular markers used in early positioning) are RFLP markers, but because such markers require complex operating procedures, expensive biochemical reagents and a large number of samples of DNA, it is not convenient for the detection of non-toxic genes. Toxic composition analysis

Method used

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  • Molecular marker detection method of rice blast bacterium non-toxic genes Avr-Pit and primers thereof
  • Molecular marker detection method of rice blast bacterium non-toxic genes Avr-Pit and primers thereof
  • Molecular marker detection method of rice blast bacterium non-toxic genes Avr-Pit and primers thereof

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Effect test

Embodiment 1

[0024] The molecular marker and location of the avirulent gene Avr-Pit linkage of Magnaporthe grisea were obtained by the following methods:

[0025]1) Using the parents Guy11 and JS153 (the parent strain is a strain preserved in the Department of Plant Diseases of Nanjing Agricultural University) and the sexual offspring obtained by crossing (identified by Chen Qinghe, etc.), the initially obtained SSR marker m355 linked to the avirulent gene Avr-Pit was inherited -356 based on (Chen QH, Wang YC, Li AN, Zhang ZG, Zheng XB.2007.Molecularmapping of two cultivar-specific avirulence genes in the rice blast fungus Magnaporthe grisea.Mol Genet Genomics.277(2):139-48 .) to design CAPS marker primers, the primer sequences are shown in Table 1:

[0026] Table 1

[0027]

[0028] 2) The molecular markers amplified by two pairs of CAPS molecular marker primers containing CAPS markers CAPS77 and CAPS88 were screened for markers linked to the avirulent gene Avr-Pit in the two parents....

Embodiment 2

[0031] The primers CAPS77 and CAPS88 in Example 1 were amplified in 65 offspring, and the PCR reaction system and program were the same as in Example 1. After the reaction was finished, 50 μl of each sample was placed on 1% agarose gel in 1×TAE ( Electrophoresis in 10mM Tris, pH 7.8, 5mM sodium acetate, 0.5mM EDTA), then soak the amplified product in a solution containing ethidium bromide (0.5μg / ml) for 10-15min, then move to clean water and rinse for 5-10min , observed under UV light and photographed with a Bio-rad gel imaging system. The fragments were recovered, digested with the corresponding enzymes EarI and PciI, and the results were observed by electrophoresis after 10 hours. If two fragments of 268bp and 101bp are produced by digestion with the enzyme EarI, the genome of the rice blast fungus to be tested contains the avirulent gene Avr-Pit; if two fragments of 472bp and 382bp are produced by digestion with the enzyme PciI The fragment of the blast pathogen to be test...

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Abstract

The invention discloses a method for rapidly detecting rice blast bacterium non-toxic genes Avr-Pit by utilizing molecular markers. In the invention, two pairs of specific primers are used, wherein an upstream primer in one pair of primers is ACCGCGATCAGTGGAAAACT, and a downstream primer is ATTGGCAGAGCCAGCTACTC; and an upstream primer in the other pair of primers is TTGTGTTCCTGTCAATCGCG, and a downstream primer is CGCAGCTTATATCTCGGATAG. The method in the invention has the advantages of simple operation, explicit result and time and cost saving.

Description

technical field [0001] The invention relates to a molecular marker primer for detecting the rice blast fungus avirulent gene Avr-Pit, and a molecular marker method for detecting the rice blast fungus avirulent gene Avr-Pit using the primer. Background technique [0002] Rice is one of the important food crops, and rice blast is one of the most widely distributed and most serious rice diseases in the world, and has become a major obstacle to high and stable rice yields (Ou S H.1980, Pathogen variability and host resistance in rice blast disease.Ann Rev Phytopathol.18:167-187.), the harm it causes is seriously threatening human food security. The control of rice blast is the same as other rice diseases, through the use of fungicides, cultivation measures and the combination of planting disease-resistant varieties, and planting disease-resistant varieties is one of the most economical and effective ways to control rice blast. However, in order to effectively carry out disease-...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12Q1/04C12N15/11
Inventor 张正光董妍涵王源超郑小波董莎萌
Owner NANJING AGRICULTURAL UNIVERSITY
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