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Method for utilizing stamen petalody to mark male sterility genes of Brassica campestris ssp.chinensis

A technology of male sterility gene and head cabbage, which is applied in the biological field, can solve the problems of low efficiency and long cycle of breeding sterile lines, and achieve the effect of optimized process, high cost and long cycle

Inactive Publication Date: 2013-03-20
NANJING AGRICULTURAL UNIVERSITY
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Problems solved by technology

[0005] The traditional breeding method has a long cycle and low efficiency for the selection of male sterile lines in cabbage, which is also an important application that currently limits the selection and application of male sterile lines in cabbage.
At present, there is no successful report on the gene differential expression analysis of the new line of non-heading cabbage with petalized stamens as a material, and then to obtain male sterility-related genes

Method used

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  • Method for utilizing stamen petalody to mark male sterility genes of Brassica campestris ssp.chinensis
  • Method for utilizing stamen petalody to mark male sterility genes of Brassica campestris ssp.chinensis

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Embodiment Construction

[0031] Implementation procedure of the present invention is:

[0032] 1. Materials and methods

[0033] The non-heading Chinese cabbage stamen petalized line W053 and its maintainer line W052, a near-isogenic line, came from the Chinese cabbage research group of the Vegetable Department of the College of Horticulture, Nanjing Agricultural University.

[0034] 2. Screening and analysis of molecular markers AFLP markers and SCAR markers were mainly used.

[0035] Use 64 pairs of selective amplification primers for PCR reaction, the system is 20 μl, including 2 μl template, 1 μl left primer, 1 μl right primer, 10 × PCR buffer (Mg 2+ free) 2μl, dNTP (2.5mM) 1.6μl, Mg 2+ 1.2μl, 5U rTaq 0.2μl, add ddH 2 0 to 20 μl; PCR reaction program: pre-denaturation at 94°C for 2 minutes, first-round amplification parameters: 94°C for 30 s, 65°C for 30 s, 72°C for 45 s, 12 rounds of cycles, with the temperature decreasing by 0.7°C in each cycle. Second-round amplification parameters: 94°C fo...

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Abstract

The invention relates to a method for utilizing stamen petalody to mark male sterility genes of Brassica campestris ssp.chinensis, belongs to the field of a biotechnology and is used for marker-assisted selection breeding of a Brassica campestris ssp.chinensis male sterility line. The method comprises the following step of: amplifying DNA (Deoxyribonucleic Acid) synthesized by RNA (Ribonucleic Acid) inverse transcription of a stamen petalody line and a maintainer line thereof by utilizing a mark primer AFLP17 (Amplified Fragment Length Polymorphism 17) or a mark primer SCAR33, wherein if a 300bp segment can be amplified, the male sterility genes of the Brassica campestris ssp.chinensis do not exist. The method can be used for carrying out cDNA-AFLP and SCAR molecular marker sieving on the DNA synthesized by the RNA inverse transcription of the Brassica campestris ssp.chinensis stamen petalody line W053 and the maintainer line W052 thereof, can effectively expand the marker-assisted selection breeding of the Brassica campestris ssp.chinensis male sterility line, and greatly improves the selection efficiency, so that the breeding progress is accelerated.

Description

1. Technical field [0001] The invention discloses a method for marking the male sterile gene of non-heading Chinese cabbage by stamen petalization, which belongs to the field of biotechnology and is used for molecular marker-assisted selection breeding of male sterile lines of non-heading Chinese cabbage. 2. Technical background [0002] In recent years, the demand for non-heading cabbage (Brassica campestris L.ssp.chinensis Makino) has been increasing, but it is not resistant to storage and transportation, and its variety is relatively single. This makes the use of heterosis more and more important, and obtains high-quality, high-yield, and stress-resistant non-heading Chinese cabbage varieties through hybridization. At present, the methods widely used to prepare hybrids mainly include self-incompatibility lines, chemical male sterilization and male sterile lines, etc., but the first two methods have great defects in terms of workload and effect respectively. Therefore, th...

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 胡春梅李树燕
Owner NANJING AGRICULTURAL UNIVERSITY
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