Method for efficiently obtaining regeneration plant by cultivating isolated microspores of brassica oleracea L.var.capitata L.

A technology for cabbage and plant regeneration, applied in the biological field, can solve problems such as embryogenesis rate and low plant regeneration frequency, and achieve the effects of quality improvement, regeneration rate improvement and mortality reduction.

Active Publication Date: 2015-03-25
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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Problems solved by technology

[0004] The purpose of the present invention is to provide a new method for efficiently obtaining regenerated plants from the free microspo...

Method used

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  • Method for efficiently obtaining regeneration plant by cultivating isolated microspores of brassica oleracea L.var.capitata L.
  • Method for efficiently obtaining regeneration plant by cultivating isolated microspores of brassica oleracea L.var.capitata L.
  • Method for efficiently obtaining regeneration plant by cultivating isolated microspores of brassica oleracea L.var.capitata L.

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Embodiment 1

[0038] (1) Embryogenesis

[0039] 1) Donor material selection Select buds of different lengths from healthy donor plants, and use tweezers to extrude some microspores. Acetate magenta or 4,6-diamididine-2-phenylindole dihydrochloride (DAPI) fluorescence After dyeing, the microspore development period was observed under a microscope, and the flower bud with the microspore in the later mononuclear stage was selected as the donor flower bud. There were differences in the length of flower buds corresponding to the optimal culture period of different genotypes, but basically the proportion of microspores in the later mononuclear stage was the highest among the flower buds in the range of 3.5-4.5 mm.

[0040] 2) Sterilization of the donor material: Wash the flower buds with purified water, then disinfect the buds with 75% alcohol by volume for 30s in an ultra-clean workbench, then disinfect the surface with a 7% volumetric sodium hypochlorite solution for 15 minutes, and then rinse ...

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Abstract

The invention discloses a method for efficiently obtaining a regeneration plant by cultivating isolated microspores of brassica oleracea L.var.capitata L.. The method comprises the following steps: (1) performing embryogenesis; and (2) performing embryo germination, thereby obtaining the regeneration plant, wherein the separated microspores are subjected to dark culture in a culture box at the temperature of 18+/-0.5 DEG C until an embryoid visible to the naked eyes is generated in the step (1) of performing embryogenesis; and then the small embryoid is transferred to a dark shaking table at a speed of 50-55 revolutions per minute at the temperature of 25+/-0.5 DEG C to perform shake culture until the cotyledonous embryoid is formed. According to the method, the genetically homogenous double-haploid plant materials can be obtained in batches, the germplasm resource innovation is accelerated, and the breeding progress is shortened. The method disclosed by the invention can be popularized and applied in the research field of molecular marker assisted breeding, mutant screening and cell totipotency mode systems of the brassica oleracea L.var.capitata L..

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a method for efficiently obtaining regenerated plants by culturing free microspores of cabbage. technical background [0002] Cabbage (Brassica oleracea L.var.capitata L.) is abbreviated as cabbage, also known as cabbage, cabbage, cabbage, cabbage, etc. in China. It is one of the most important vegetable crops in the Brassica family. Cabbage is a cross-pollinated plant with obvious heterosis. However, as a chloroplast vernal plant, limited by conditions, generally only one generation of self-propagation can be propagated a year, and it takes at least 5 to 6 years to develop a stable inbred line. Free microspore culture is one of the best methods to quickly obtain homozygous double haploids, which plays an important role in accelerating the selection of new crop varieties and improving the breeding efficiency. In addition, episomal microspore culture can also be applied to the researc...

Claims

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Application Information

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IPC IPC(8): A01H4/00
Inventor 曾爱松严继勇宋立晓高兵
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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