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Medium and culture method for indirect regeneration of highbush blueberry somatic embryos

A highbush blueberry and somatic embryo technology, applied in biochemical equipment and methods, plant regeneration, horticultural methods, etc., can solve the problem of failure to obtain embryogenic callus, no rooting medium formula for buds, and no obvious application Value and other issues to achieve the effect of reducing the risk of browning, reducing the accumulation of harmful substances, and ensuring the supply of nutrients

Active Publication Date: 2018-11-27
WENZHOU VOCATIONAL COLLEGE OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The success of this application lies in the establishment of a method for direct induction of embryoid bodies from blueberry stem tips; but the disadvantages are that the proliferative embryogenic callus cannot be obtained without the process of callus dedifferentiation into seedlings, There is no medium formula for shoots and roots, so it has no obvious application value in molecular breeding, especially in subsequent cell culture and protoplast culture

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  • Medium and culture method for indirect regeneration of highbush blueberry somatic embryos
  • Medium and culture method for indirect regeneration of highbush blueberry somatic embryos
  • Medium and culture method for indirect regeneration of highbush blueberry somatic embryos

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preparation example Construction

[0054] 1. Preparation of four culture media

[0055] The medium used for the indirect regeneration of highbush blueberry somatic embryos of the present invention includes four culture media for different culture stages in the highbush blueberry somatic embryo indirect regeneration, which are respectively: (1) highbush blueberry callus Tissue carbon starvation liquid induction medium, (2) highbush blueberry embryogenic callus induction and proliferation semi-solidified medium, (3) highbush blueberry somatic embryo maturation and differentiation, organic medium for seedling regeneration, (4) ) rooting medium of highbush blueberry differentiated seedlings.

[0056] Examples of the preparation of the four media are as follows:

Embodiment 1

[0058] (1) Highbush blueberry callus carbon starvation liquid induction medium: add ZT0.5mg / L, TDZ0.1mg / L, 2-ip 0.25mg / L and CPPU 0.75mg / L to the improved 1 / 2MS basic medium L, continue to add sucrose 5g / L, do not add agar, adjust the pH to 5.0;

[0059] (2) Highbush blueberry embryogenic callus induction and proliferation semi-solidified medium: add ZT 1mg / L, TDZ0.2mg / L, 2-ip 0.5mg / L and CPPU 1.5 to the improved 1 / 2MS basic medium mg / L, continue to add sucrose 30g / L, agar 3g / L, and adjust the pH to 5.0;

[0060] (3) Organic medium for maturation, differentiation and seedling regeneration of highbush blueberry somatic embryos: add ZT 1mg / L and IBA 0.1mg / L to the improved 1 / 2MS basic medium, continue to add sucrose 20g / L, coconut Milk 100mg / L, AC 0.5g / L, agar 5g / L, adjust pH to 5.0;

[0061] (4) Rooting medium for highbush blueberry differentiated seedlings: add IBA 1.0mg / L, sucrose 3.5g / L, AC 0.5g / L to the improved 1 / 6MS basic medium, without adding agar, and adjust the pH t...

Embodiment 2

[0063] (1) Highbush blueberry callus carbon starvation liquid induction medium: add ZT 0.7mg / L, TDZ0.15mg / L, sucrose 8g / L, all the other are identical with embodiment 1;

[0064] (2) Highbush blueberry embryogenic callus induction and proliferation semi-solidified medium: add agar 4g / L, and the rest are the same as in Example 1;

[0065] (3) The maturation and differentiation of highbush blueberry somatic embryos, the organic matter medium of seedling regeneration: add ZT 1.6mg / L agar 6g / L, all the other are identical with embodiment 1;

[0066] (4) rooting medium of highbush blueberry differentiated seedling: same as embodiment 1;

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Abstract

The invention relates to the field of tissue culture of plants, aims to provide a culture medium for indirect regeneration of high-bush blueberry somatic embryos and a culture method. The method includes the steps: preparing four culture media; taking stems with leaf stalks of sterile test-tube plantlets of high-bush blueberries under sterile operation conditions; performing low-temperature and low-light shallow liquid culture for calluses, and performing low-temperature and dark differentiation multiplication culture for semi-solidification culture media embryonic calluses of blueberries; performing full-light and normal-temperature differentiation seedling culture for solid media of the blueberry embryos, and performing rooting seeding culture. According to the method, multiplication speed of non-embryonic calluses is reduced in the induction process of the calluses, and a browning risk caused by high temperature and long-term non-subculture is reduced. Four types of cytokinin are matched in low concentration, callus states of embryonic calluses are effectively maintained, multiplication of the embryonic calluses after successful induction is facilitated, the culture media and other media cannot be adhered on rooting seedlings, transplanting efficiency is high, and labor cost is remarkably reduced.

Description

technical field [0001] The invention relates to a method for high-frequency induction and proliferation of embryogenic callus of blueberries and a somatic embryo differentiation into seedlings, belonging to the field of plant tissue culture. Background technique [0002] A callus is a group of dedifferentiated cells that undergo cell division to produce a disorganized, non-polar, loose cell mass. According to histological observations, appearance characteristics, regeneration, and regeneration methods, callus can be divided into two categories: embryogenic callus (EC) and non-embryonic callus (NEC, NEC). Such as figure 1 ). Generally, the embryogenic callus is solid in texture, milky white or yellow in color, with spherical particles on the surface, and its growth is slow; from the perspective of cytology, the embryogenic callus is composed of cells of equal diameter, the cells are small, the protoplasm is thick, and there is no Vacuoles, often rich in starch grains, with...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/00A01H4/00
CPCA01H4/001A01H4/008
Inventor 余宏傲王法格叶朝军金微微康华靖
Owner WENZHOU VOCATIONAL COLLEGE OF SCI & TECH