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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  • Abstract
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  • Application Information

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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 bacteri

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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Example Embodiment

[0037] Example 1:

[0038] 1. Material preparation

[0039] 1. Plant materials

[0040] Strawberry "Hongyan" tissue cultured seedlings were subcultured on strawberry subculture medium for experiment.

[0041] 2. Medium formula and preparation method:

[0042] Strawberry ‘Hongyan’ Subculture Medium FMS1:

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

[0044] Strawberry ‘Hongyan’ co-cultivation medium FMS2:

[0045] MS4.4g / L, sucrose 30g / L, agar 5.5 g / L, TDZ 2mg / L, 2,4-D 0.1mg / L, pH 5.8, autoclaved, cooled to about 60℃ and packed into flat plates for later use;

[0046] Strawberry ‘Hongyan’ Screening 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, autoclaved, cooled to 60℃ and added kanamycin 0 -50mg / L, carbenicillin 100-400 mg / L sub-packed culture flasks for use;

[0048] Strawberry ‘Hongyan’ Rooting Screening Medium FMS4:

[0049] MS4.4g / L, sucrose 30g / L, agar 5.5 g / L, IBA 0.1mg / L, pH5.8, autocla...

Example Embodiment

[0092] Example 2: Basically the same as Example 1, except that: Strawberry "Hongyan" efficient, rapid and stable gene transformation method is as follows:

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

[0094] Step 3 Infection process: Pour the suspended bacterial liquid into the cut leaves, shake it gently, soak for 60 minutes, and shake every 5 minutes during the process to increase the contact area between the bacterial liquid and the leaves, and pour after the infection is complete Bacteria liquid is discharged, and the leaves are dried on sterile filter paper to dry the bacterial liquid on the surface.

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

[0096] Step 5 Screening the cultiva...

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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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