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Dry-germ dissociation method applied to wheat mature embryo culture

A wheat and grass plant technology, applied in the field of dry embryo dissociation, can solve the problems of time-consuming and labor-intensive, easy to pollute, complicated operation, etc., and achieve the effects of saving storage space, labor, and facilitating tissue culture and genetic transformation.

Inactive Publication Date: 2013-12-11
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The inoculation methods of the above predecessors all sterilize the seeds, and then peel and inoculate the embryos soaked overnight in the ultra-clean bench. The operation is cumbersome, easy to pollute, and time-consuming and labor-intensive.

Method used

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  • Dry-germ dissociation method applied to wheat mature embryo culture
  • Dry-germ dissociation method applied to wheat mature embryo culture
  • Dry-germ dissociation method applied to wheat mature embryo culture

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Embodiment 1, the exploration of experimental conditions

[0045] 1. Preparation of dry embryo tissue

[0046] Evenly crush the grains of the wheat variety "Liaochun No. 18", first pass through a 9-mesh sieve to remove large particles, and then pass through a 30-mesh sieve to remove small particles, from the particles whose particle size is between 9-mesh sieve and 30-mesh sieve Pick embryonic tissue (light yellow, crescent-like in shape, 1-2mm in size, accounting for about 40% of the broken tissue), and put it in a 1.5ml centrifuge tube for later use. For photos during the screening see figure 1 , A and B are particles whose particle size is between 9 mesh sieve and 30 mesh sieve, C is endosperm particle whose particle size is between 9 mesh sieve and 30 mesh sieve, D is between 9 mesh sieve and 30 mesh sieve The granules of the embryo in between.

[0047] 2. Exploration of disinfection time

[0048] 1. Take the embryo tissue obtained in step 1, wash it 2-3 times w...

Embodiment 2

[0065] Embodiment 2, the application of the method provided by the present invention

[0066] 1. Evenly crush the grains of the wheat variety "Liaochun No. 18", first pass through a 9-mesh sieve to remove large particles, and then pass through a 30-mesh sieve to remove small particles, from the grain size between 9-mesh sieve and 30-mesh sieve Pick the embryonic tissue (light yellow, crescent-shaped, 1-2mm in size, accounting for about 40% of the broken tissue) from the granules, and put it in a 1.5ml centrifuge tube for later use.

[0067] 2. Take the embryo tissue obtained in step 1, wash it with sterile water for 2-3 times, then disinfect it with 2.5g / 100ml sodium hypochlorite aqueous solution for 1 minute, and then rinse it with sterile water for 3-5 times.

[0068] 3. Inoculate the embryo tissue after step 2 into the induction medium (80 embryo tissues per culture dish, repeat 3 culture dishes), culture in the dark at 25°C for 1 week (see photo image 3 A). The callus g...

Embodiment 3

[0074] Embodiment 3, comparison method

[0075] 1. Take the grains of the wheat variety "Liaochun No. 18", soak them in 70% (volume ratio) alcohol solution for 10 minutes, and then soak them in 2.5g / 100ml sodium hypochlorite solution for 25 minutes.

[0076] 2. Take the seeds that have completed step 1 and soak them in water for 12 hours.

[0077] 3. Take the seeds that have completed step 2 and soak them in sodium hypochlorite solution for 15 minutes.

[0078] 4. Take the seeds that have completed step 3, scrape the embryos of the seeds gently with a sharp knife 5-6 times, and inoculate the mince (the mince of each embryo is inoculated to the same position of the induction medium as 1 piece) into the induction medium , cultured in the dark at 25°C for 1 week (see image 3 B). The callus generation rate was 94.1% (the average value of 3 petri dishes).

[0079] 5. Same as step 4 of embodiment 2.

[0080] 6. Same as step 5 of embodiment 2. The callus was cultured in the di...

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Abstract

The invention discloses a dry-germ dissociation method applied to wheat mature embryo culture, and provides an embryo taking-out method of grasses. The method comprises the following steps: smashing kernels of the grasses, using a 9-mesh mesh sieve to remove large particles, then using a 30-mesh mesh sieve to remove small particles, and selecting 1-2 mm embryo tissues from particles of which the sizes are within the range of 9-30 meshes. Compared with the conventional embryo scraping method, the embryo taking-out method provided by the invention has the advantages that although difference between wound curing induction and differentiation effect in follow-up tests is insignificant, processes of embryo taking-out, storage and disinfection become convenient and fast, so that the manpower is saved; the method can be applied to the storage of germ plasm resources to save storage space. According to the method, tissue culture and genetic transformation of mature embryos on the grasses are greatly facilitated. In summary, the method has an important application significance.

Description

technical field [0001] The invention relates to a method for freeing dry embryos applied to the cultivation of mature wheat embryos. Background technique [0002] Wheat is an important food crop in my country, which is closely related to national food security, national economic development, social stability and improvement of people's living standards. In recent years, the rapid development of agricultural biotechnology has provided a new impetus for the study of wheat genetics and breeding, among which wheat transgenic technology has opened up broad prospects for the development of wheat genome research and genetic improvement. [0003] The most important foundation of wheat transgenic technology is the establishment and improvement of wheat tissue culture system. The explant sources of the wheat tissue culture system mainly include immature embryos, anthers, apical meristems, young ears, mature embryos, etc. Among them, the callus induction and regeneration ability of wh...

Claims

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Application Information

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IPC IPC(8): C12N5/04A01H4/00A01H5/00
Inventor 解超杰孙其信倪中福梁荣奇李映辉沈红霞彭福祥宋娜耿妙苗
Owner CHINA AGRI UNIV
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