Efficient, rapid and stable genetic transformation method for strawberries

A technology for gene transformation and strawberry, applied in the field of genetic engineering, can solve the problems of large workload of transgenic detection, contamination of Agrobacterium and miscellaneous bacteria, low efficiency of strawberry transformation, etc., so as to avoid Agrobacterium contamination, reduce damage and improve transgenic efficiency. Effect

Active Publication Date: 2014-12-10
JIANGSU POLYTECHNIC COLLEGE OF AGRI & FORESTRY
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AI Technical Summary

Problems solved by technology

[0005] In order to solve the above-mentioned technical problems, the present invention provides a method for efficient, rapid and stable gene transformation of strawberry 'Hongyan', which not only solves the problem of contamination of Agrobacterium and miscellaneous bacteria in the traditional Agrobacterium-mediated method, and the problem of low transformation efficiency of strawberry; Transgenic from transgenic, transgenic detection, long transplanting period in the field, the workload of transgenic detection is heavy, and the work is heavy

Method used

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  • Efficient, rapid and stable genetic transformation method for strawberries
  • Efficient, rapid and stable genetic transformation method for strawberries
  • Efficient, rapid and stable genetic transformation method for strawberries

Examples

Experimental program
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Effect test

Embodiment 1

[0038] 1. Material preparation

[0039] 1. Plant material

[0040] Strawberry "Hongyan" tissue culture seedlings were subcultured on the strawberry subculture medium for experimentation.

[0041] 2. Culture medium formula and preparation method:

[0042] Strawberry 'Hongyan' subculture medium FMS1:

[0043] MS4.4g / L, sucrose 30g / L, agar 5.5 g / L, 6-BA0.5mg / L, NAA0.1mg / L, pH5.8, autoclaved, ready for use;

[0044] Strawberry 'Hongyan' co-culture medium FMS2:

[0045] MS 4.4g / L, sucrose 30g / L, agar 5.5 g / L, TDZ 2mg / L, 2,4-D 0.1mg / L, pH 5.8, autoclave, cool to about 60°C and pack into plates for later use;

[0046] Strawberry 'Hongyan' selection medium FMS3:

[0047] MS4.4g / L, sucrose 30g / L, agar 5.5 g / L, TDZ 2mg / L, 2,4-D 0.1mg / L, pH5.8, autoclave, cool to 60°C and add kanamycin 0 -50mg / L, carbenicillin 100-400 mg / L subpackage culture bottle for later use;

[0048] Strawberry 'Hongyan' Rooting Screening Medium FMS4:

[0049] MS4.4g / L, sucrose 30g / L, agar 5.5 g / L, IBA 0.1mg / L...

Embodiment 2

[0092] Embodiment 2: basically the same as Example 1, the difference is: the efficient, rapid and stable gene transformation method of strawberry 'Hongyan', specifically as follows:

[0093] In the second step of step 1 strain activation, pick 2-3 good single colonies that grow on the plate, inoculate them into 50 mL LB liquid medium, shake and cultivate in a shaker at 28 ℃, 220 rpm until the OD600 value 0.6.

[0094] Step 3 Infection process: Pour the suspended bacterial solution into the cut leaves, shake slightly, and dip for 60 minutes. During the process, shake once every 5 minutes to increase the contact area between the bacterial solution and the leaves. Bacterial solution was removed, and the leaves were blotted dry on the sterile filter paper to dry the bacterial solution on the surface.

[0095] Step 4 Co-cultivation process: the infected leaves are inoculated in the strawberry co-culture medium FMS2, and cultured in the dark for 2 days in an incubator at 25±2°C.

...

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Abstract

The invention discloses an efficient, rapid and stable genetic transformation method for strawberries 'beauty'. The method comprises the steps of activation of culture, preparation of plant materials, infection, co-culture, screening culture and rooting culture, solves the problems of the pollution of agrobacterium and infectious microbes in a traditional agrobacterium-mediated method, low conversion efficiency of the strawberries 'beauty' and long period of the strawberries 'beauty' from transgenosis and transgenic detection to field transplanting, shortens the period, further solves the problems of large workload and heavy work of transgenic detection, simplifies a detection method, and increases the detection speed. On the basis of the established 'beauty' strawberry leaf regeneration system, the method researches various factors for influencing agrobacterium tumefaciens transforming the 'beauty' strawberry leaves, optimizes various parameters, simplifies the operation process, and determines the best transformation conditions.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a strawberry transgenic technology. Background technique [0002] strawberry( Fragaria ananassa ) is a crop with high economic value and high nutritional value. In recent years, the strawberry industry has developed rapidly in many areas of our country, and has become an important part of agricultural industrialization in some areas. Therefore, in recent years, Chinese scientific researchers have also invested a lot of energy in the research of strawberry, especially with the rapid development of molecular biology and genetic engineering in recent years, the research focus has turned to the deep excavation of excellent genes of strawberry germplasm resources, and carried out Identification and determination of molecular markers and gene expression sequences. At the same time, research results on genetic improvement such as stress resistance, storage and ...

Claims

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Application Information

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IPC IPC(8): C12N15/84A01H5/00
Inventor 王媛花蔡善亚颜志明董慧杨宝林
Owner JIANGSU POLYTECHNIC COLLEGE OF AGRI & FORESTRY
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