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Method for identifying paddy female sterile gene FST molecule

A technology for molecular identification and femaleness, which is applied in biochemical equipment and methods, and microbial measurement/inspection, etc., can solve the problems of blocked development of ovule megaspores, lack of embryo sac, cumbersome methods, etc., and achieve a simple and easy identification method, Avoid cumbersome procedures, simple and economical identification method

Inactive Publication Date: 2012-08-22
YUNNAN AGRICULTURAL UNIVERSITY
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AI Technical Summary

Problems solved by technology

②Although it has female organs (with or without style and stigma), the development of ovules and even the development of megaspores is hindered and there is no normal embryo sac
These two detection methods need to plant the materials, and the observation or hybridization can only be done when the ovules of the plants develop. The time period is about 90 days, and the method is cumbersome.
Therefore, there is not yet a method with perfect technology, fast and effective selection and identification of the female sterility gene FST

Method used

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  • Method for identifying paddy female sterile gene FST molecule
  • Method for identifying paddy female sterile gene FST molecule
  • Method for identifying paddy female sterile gene FST molecule

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] From the Ansanbyeo / G39 hybrid F2 population, a total of 220 individual plants, the genotype of the 220 individual plants was identified by embryomorphological observation and molecular identification method, and the identification results of the two methods were compared to investigate the reliability of the molecular identification method .

[0025] When observing embryo morphology, the embryo sac after flowering was stripped, fixed with glutaraldehyde fixative (1.4% glutaraldehyde, 2% paraformaldehyde, 50mM PIPES, pH 7.20), rinsed with PIPES, dehydrated with ethanol step by step (10 %-100%), embedded in paraffin, sectioned (2um), stained (0.5% toluidine blue + 0.1% Na 2 CO 3 ), examined under an optical microscope.

[0026] For molecular identification, take the leaves of the materials to be identified and extract the total DNA of rice; use FST-specific primers Gf1, Gr1, Jf1 and Jr1 for PCR reaction; add 3 μl of loading buffer, mix well, load 6 μl of the sample, and...

Embodiment 2

[0040] From Sangambyeo / G39 hybrid F2 generation population, a total of 340 individual plants were used to identify the genotypes of the 340 individual plants by hybridization verification and molecular identification methods. The identification results of the two methods were compared to investigate the reliability of the molecular identification method.

[0041] During hybridization verification, when paddy rice blooms, get fresh pollen to pollinate its stigma, bagging. Each plant was bagged with one ear, and the fruiting situation was inspected in two weeks.

[0042] For molecular identification, take the leaves of the material to be identified and extract the total DNA of rice; use FST specific primers Gf1, Gr1, Jf1 and Jr1 for PCR reaction; add 3 μl of loading buffer to mix and load 6 μl of the sample, and use 2% agarose gel, 1×TAE buffer, 125V electrophoresis for 40min; detect on the ultraviolet transilluminator, and identify the genotype of the material according to the ...

Embodiment 3

[0056] 20 rice male sterile lines and 50 various rice resources, 70 materials are all female fertile materials, the genotype of the materials was identified by molecular identification method, and the reliability of molecular identification method was investigated.

[0057] For molecular identification, take the leaves of the materials to be identified and extract the total DNA of rice; use FST-specific primers Gf1, Gr1, Jf1 and Jr1 for PCR reaction; add 3 μl of loading buffer, mix well, load 6 μl of the sample, and use 2% agarose gel , 1×TAE buffer, 125V electrophoresis for 40 minutes; detect on the ultraviolet transilluminator, and identify the genotype of the material according to the electrophoresis results. The molecular weights of PCR products whose genotype is wild type (+ / +) are 395bp and 663bp; the molecular weights of PCR products whose genotype is mutant (- / -) are 298bp and 655bp, and the mutant type is female sterile material; the genotype is heterozygous The molec...

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Abstract

The invention relates to a method for identifying paddy female sterile gene FST molecules, which belongs to the breeding field of agricultural biological technology. The adopted technical scheme comprises the following steps: extracting paddy total DNA, and carrying out PCR reaction by particularity primers Gf1, Gr1,Jf1 and Jr1; adding 3 mu l of loading buffer to be uniformly mixed, sampling 6 mul, and carrying out the electrophoresis of 125 V for 40 minutes by agarose gel of 2 percent and 1*TAE buffer; and checking on an ultraviolet transilluminator, and identifying the gene type of the material according to the electrophoresis result. The gene type is a wild PCR product with the molecular weight of 395bp and 655bp, the PCR product of a mutation type (female sterile) has the molecular weight of 298 bp and 655 bp, and the PCR product of a hybridization type has the molecular weight of 298 bp, 395 bp and 663 bp (or 655bp). The invention has the effect of rapidly and effectively identifying the paddy female sterile gene FST.

Description

technical field [0001] The invention relates to a method for molecular identification of rice female sterility gene FST, which belongs to the field of agricultural biotechnology, and more specifically belongs to the field of breeding. Background technique [0002] The breakthrough of crop breeding depends on the discovery of new specific germplasm materials. The most prominent example in rice is the discovery and utilization of cytoplasmic male sterile lines in the 1970s. Male sterility is the period from the formation of stamen primordia to the formation of functional mature pollen grains. The process of physiological, biochemical, and morphological changes is blocked, and stamens cannot develop normally, resulting in the inability to form viable pollen. The phenomenon. [0003] The opposite of male sterility is female sterility. Normal plant female organ development mainly refers to the development of the ovary (including ovule development and embryo sac formation). If t...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 李东宣陈丽娟李成云李娟甘树仙谭亚玲刘永胜徐学洙朱有勇
Owner YUNNAN AGRICULTURAL UNIVERSITY
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