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Method for culturing regeneration plants of Brassica oleracea microspores

A technology for regenerating plants and cultivation methods, applied in the field of plant tissue culture, can solve problems affecting the quality of embryos, affecting the embryo production rate, inhibiting microspore cultivation and breeding technology, etc., and achieve the effect of improving utilization efficiency and embryo emergence rate

Inactive Publication Date: 2012-11-07
ZHEJIANG UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

A large number of studies on the culture of free microspores of Brassica have shown that the genotype of the donor plant is extremely important for the microspore embryogenesis, which not only affects the embryo production rate, but also affects the quality of the embryo (Chuong et al., 1988) , inhibiting the application of microspore culture breeding technology

Method used

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  • Method for culturing regeneration plants of Brassica oleracea microspores

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] The cultivation method is carried out as follows.

[0036] (1) Culture medium preparation: comprise the culture medium of microspore different culture stages, its component and the weight that each component contains in every liter of medium are:

[0037] Table 1 NLN and MS medium formula table

[0038]

[0039] 1) NLN-13 induction medium: NLN liquid medium 1L + sucrose 130g / L, pH6.0, filter sterilized;

[0040] 2) Embryoid body differentiation medium: MS medium + sucrose 20g / L, agar 10g / L, pH 6.0, high temperature and high humidity sterilization;

[0041] 3) Rooting medium: MS medium + sucrose 30g / L, agar 6g / L, pH5.8, high temperature and high humidity sterilization;

[0042] (2) The cultivation of the microspore regenerated plant of broccoli with high germination rate:

[0043] 1) Selection of flower buds of donor plants: take inflorescences of broccoli and rapeseed that are healthy and free from diseases and insect pests as donor plants for microspore culture; ...

Embodiment 2

[0053] In addition to step (1) in 1) NLN-13 induction medium: NLN-13 liquid medium 1L + sucrose 130g / L, pH6.1, filter sterilization; 2) embryoid differentiation medium: MS medium + sucrose 20g / L, agar 11g / L, pH 6.1, high temperature and high humidity sterilization; 3) rooting medium: MS medium + white sugar 20g / L, agar 7g / L, pH 5.9, high temperature and high humidity sterilization;

[0054] (2) The cultivation of the microspore regenerated plant of broccoli with high germination rate:

[0055] 1) Selection of flower buds of donor plants: take inflorescences of broccoli and rapeseed that are healthy and free from diseases and insect pests as donor plants for microspore culture; 1.2. Flower buds from late mononucleate to early binucleate;

[0056] 2) Sterilization of flower buds: use 5.6% (mass percentage concentration) sodium hypochlorite aqueous solution 56ml / L+ absolute ethanol 100ml / L+ detergent 8 drops+ sterile water to prepare the sterilizing solution; mix rape and brocc...

Embodiment 3

[0065] In addition to step (1) 1) NLN-13 induction medium: NLN-13 liquid medium 1L + sucrose 130g / L, pH 6.2, filtered and sterilized; 2) Embryoid body differentiation medium: MS medium + sucrose 20g / L, agar 12g / L, pH6.0, high temperature and high humidity sterilization; 3) rooting medium: MS medium + sucrose 25g / L, agar 6.5g / L, pH5.6, high temperature and high humidity sterilization;

[0066] (2) The cultivation of the microspore regenerated plant of broccoli with high germination rate:

[0067] 1) Selection of flower buds of donor plants: take the inflorescences of broccoli and rape that are healthy and free of diseases and insect pests as donor plants for microspore culture; take flower buds with a petal to anther length ratio of 0.9 on the inflorescence, late mononucleate to early binucleate;

[0068]2) Sterilization of flower buds: use 5.6% (mass percentage concentration) sodium hypochlorite aqueous solution 56ml / L+dehydrated alcohol 100ml / L+clean liquid 9 drops+sterile w...

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Abstract

The invention discloses a method for culturing regeneration plants of Brassica oleracea microspores, which comprises the following steps of: taking flower buds from inflorescences of Brassica oleracea and Brassica napus directly or taking the flower buds from the inflorescences of the Brassica oleracea and the Brassica napus after inducing; sterilizing, adding an induced culture medium NLN-13 to prepare a suspension, filtering to obtain filter liquor, and centrifuging to obtain precipitates; adding the induced culture medium NLN-13 and mixed solution of active carbon sequentially to obtain a microspore suspension; performing heat activation, and culturing to obtain embryoid in the cotyledon stage; inoculating to an embryoid differential medium until the embryoid is differentiated to germinate; cutting regenerated sprouts, inoculating to a rooting culture medium to perform rooting culture, and hardening seedlings and transplanting to obtain the regeneration plants; and taking tender leaves of the regeneration plants to detect the ploidy of the regeneration plants, and identifying regenerated seedlings of the Brassica napus and regenerated seedlings of the Brassica oleracea from a regeneration plant colony. In the method, the Brassica napus of which the embryos are easy to germinate and the Brassica oleracea of which the embryos are difficult to germinate are mixed to be cultured, and materials of which the embryos are easy to germinate drives materials of which the embryos are difficult to germinate to germinate the embryos, so that the embryo growth rate of the Brassica oleracea is improved.

Description

technical field [0001] The invention relates to the technical field of plant tissue culture, in particular to a method for cultivating broccoli microspore regeneration plants. Background technique [0002] Broccoli (B rassica oleracea L var. italica), also known as broccoli, green cauliflower, stem broccoli, etc., is a primary or secondary herbaceous plant with green curd as the product organ in Brassicaceae Brassica species. The green flower ball formed on the top of the main stem and side branches is used as the product. It is rich in nutrition, good in color, aroma and taste, and is a famous and special vegetable that is very popular in the international market. At present, most of the main varieties of broccoli in my country come from abroad, and the varieties with our independent intellectual property rights are scarce. The reason is that our own broccoli breeding resources are few and we cannot prepare good materials. The method is to self-segregate the varieties used ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01H4/00
Inventor 张振超周伟军庄义庆许玲耿鑫鑫
Owner ZHEJIANG UNIV
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