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A method for removing apple virus by ultra-low temperature vitrification of shoot tips

A vitrification ultra-low temperature, apple virus technology, applied in horticultural methods, botanical equipment and methods, plant regeneration and other directions, to achieve the effects of simple operation, good technical effects and promotion prospects, and improved detoxification efficiency

Active Publication Date: 2020-12-01
NANJING AGRICULTURAL UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the ultra-low temperature detoxification technology is not yet complete, and the detoxification integration process needs to be further optimized, so as to maximize the detoxification efficiency, reduce costs, and provide people with a convenient, practical, simple and efficient detoxification method

Method used

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  • A method for removing apple virus by ultra-low temperature vitrification of shoot tips
  • A method for removing apple virus by ultra-low temperature vitrification of shoot tips
  • A method for removing apple virus by ultra-low temperature vitrification of shoot tips

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Experimental program
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Effect test

Embodiment 1

[0074] 1. Total RNA extraction, primer design and RT-PCR virus detection of virus-carrying materials:

[0075] Four kinds of apples and rootstocks, 'Yanfu 3', 'Yanfu 10', 'M9' and 'M9T337' were used as materials in the experiment, all of which were obtained from Xuzhou Fruit Research Institute of Jiangsu Province. According to Zhao Lingling et al.’s description of the apparent symptoms of five major apple virus diseases: ACLSV, ASGV, ASPV, ApMV, and ASSVd (Zhao Lingling, Song Laiqing, Liu Meiying, Atanas Blagov, Tang Yan, Sun Yanxia, ​​Jiang Zhongwu. 2013. The main symptoms of apple virus diseases, Hazard and transmission route analysis. Yantai Fruit Trees, (03):5-6), selected 'Yanfu 3', 'M9', 'M9T337' plants with corresponding symptoms of poisoning and 'Yanfu 10' without symptoms No.', total RNA was extracted from its leaves, and cDNA was obtained by reverse transcription. According to the five viral protein coat sequences of ACLSV, ASGV, ASPV, APMV, and ASSVd registered in ...

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Abstract

The invention discloses a method for ultralow temperature apple virus removal by shoot tip vitrification. The method includes the following steps: a step (1) of obtaining a sterile tissue culture seedling carrying a virus; a step (2) of performing subculture and propagation; a step (3) of performing shoot tip vitrification ultralow temperature treatment for virus removal: taking the tissue cultureseedling having undergone subculture and propagation, cutting and taking a shoot tip, and inoculating the shoot tip on a pre-culture medium for pre-culture; placing the pre-cultured shoot tip in a loading solution, and loading for 5-120 min at 20-25 DEG C; performing transferring to a PVC-2 solution, performing PVC-2 vitrification for 5-150 min at 0-2 DEG C; performing liquid nitrogen refrigeration; finally performing water bath unfreezing; placing the unfreezed shoot tip in a MS + 1.2 mol / L sucrose solution for soaking; a step (4) of performing later-period proliferation propagation. The method has the advantages that (1) the virus removal efficiency is greatly improved, and the operation is simple, quick and effective; (2) an efficient technical system of the whole detailed process frommaterial selection to the obtaining of a virus-free shoot tip regeneration plant is provided; and (3) the method has a very good technical effect and promotion prospects.

Description

technical field [0001] The invention belongs to the technical field of virus-free seedling cultivation of fruit trees, and in particular relates to a method for removing apple viruses by vitrification of stem tips at ultra-low temperature. It may also have a better removal effect. Background technique [0002] Apple (Malus pumila Mill.) is a perennial deciduous tree belonging to the Malus subfamily of Rosaceae, native to Europe and central Asia, and is one of the four largest fruits in the world. Apple is rich in nutrients such as minerals and vitamins, and is a low-calorie fruit. The nutrients are highly soluble and easy to be absorbed by the body. It is also a healthy product. There are thousands of varieties of apples. There are 93 countries and regions producing apples in the world. my country is the world's largest apple producer. [0003] Apples cultivated in my country are generally infected with various viruses. In terms of production, apple virus disease is one of...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01H4/00
CPCA01H4/001A01H4/008
Inventor 高志红李艳林渠慎春栾雨婷
Owner NANJING AGRICULTURAL UNIVERSITY
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