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PCR primers and kit for screen of cabbage recessive nuclear gene male sterility, and application thereof

A technology of male sterility and nuclear gene, applied in the biological field, can solve the problem that recessive sterility is not actually used, and achieve the effects of improving breeding efficiency, simple and easy operation, and shortening breeding cycle.

Inactive Publication Date: 2019-04-02
INST OF VEGETABLE & FLOWERS CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, dominant male sterile materials have been successfully applied in cabbage breeding, while recessive sterility has not been practically utilized, and has great application potential

Method used

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  • PCR primers and kit for screen of cabbage recessive nuclear gene male sterility, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1. Development of markers and specific primers for assisted screening of cabbage recessive nuclear gene male sterility:

[0033] Resequence the whole genome of the parents "01-20" and "51S" of the cabbage recessive male sterile material "2051S" to find the SNP and Indel difference sites in the website http: / / 10.122.68.27 / cgi- bin / gbrowse / Oleracea / Find and screen the DNA sequence at the site of the difference, and then on the website http: / / bioinfo.ut.ee / primer3-0.4.0 / Design primers by DNA sequence of differential sites , use the parental DNA to screen all the primers designed to find primers with differences; then use the screened primers with differences to establish linkage markers for the mixed pool of F2 generation materials, find and determine candidate genes, according to the gene sequence in the parent The following primers were designed for the difference between "01-20" and "51s":

[0034] Upstream primer ms3-F: 5'-GAATGTATCAGGCGTGGAGG-3'

[0035]...

Embodiment 2

[0036] Embodiment 2. Kit and its preparation

[0037] The primers obtained in Example 1 were artificially synthesized, directly loaded into a centrifuge tube, or prepared into a working concentration of 10-100 times and loaded into a centrifuge tube, and then loaded into a kit.

[0038] Load the reagent containers for PCR and / or electrophoresis reagents into the kit.

Embodiment 3

[0039] Example 3. Accuracy verification of PCR primers or kits for screening the male sterility of cabbage recessive nuclear genes according to the present invention

[0040] Reagents: 2×EasyTaqPCRSuperMix purchased from Beijing Quanshijin Biology

[0041] Step 1. Extract Genomic DNA

[0042] The cabbage "01-20", "Xiaqiang", "Yafei", "Russian 05-17", "Hubei 1180", "Kag91", "308 Dongsheng", "Zhonggan 21", "Zhonggan 23", "Zhonggan 25", "Zhonggan 196", "Huifeng 4", "Huifeng 5", "Jiao "Sai", "American Purple Cabbage", "Keppel", "87-534", and the genome DNA of cabbage recessive male sterile material "2051S".

[0043] For the DNA extraction method, see Murray MG, Thompson WF (1980) Rapid isolation of high molecularweight plant DNA. Nucleic Acids Res 8:4321-4325.

[0044] Step 2, identify the amplification of primer ms3-F / ms3-R in cabbage material

[0045]In order to identify the amplification of the primers in cabbage, BGI was entrusted to synthesize primers ms3-F and ms3-R. Us...

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PUM

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Abstract

The invention discloses a molecular marker and special primers for assisting identification of cabbage recessive nuclear gene male sterility, and application thereof, and relates to molecular biological breeding technique. The name of primers for detecting the PCR marker for identifying the cabbage recessive nuclear gene male sterility is ms3-F / ms3-R; and identification of the cabbage male sterility by a reagent (primer pair) provided by the invention 100% coincides with field identification. The reagent, molecular marker or method are used for cabbage breeding, and have the advantages of simple operation, high specificity, good stability, and early breeding, and has great application prospects.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a molecular marker for assisting identification of male sterility of cabbage recessive nuclear gene, special primers and application thereof. Background technique [0002] Heading cabbage, referred to as cabbage, belongs to the Brassica cabbage species of the Cruciferae (Cruciferae). Cabbage has strong adaptability and stress resistance, and has high nutritional and health value, so it is widely cultivated in China and even all over the world. Due to the strong heterosis of cabbage, most of the excellent varieties of cabbage at home and abroad are first-generation hybrids. At present, domestic and foreign cabbage hybrids are all made of self-incompatible lines and male sterile lines. The use of self-incompatibility lines to prepare first-generation hybrids of cabbage has many disadvantages, such as the high cost of pollination and reproduction of the parents at the artificial bud st...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13
Inventor 吕红豪袁凯文韩风庆方智远杨丽梅庄木张扬勇王勇
Owner INST OF VEGETABLE & FLOWERS CHINESE ACAD OF AGRI SCI
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