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Method for improving conversion rate of foreign gene of stem tip meristem of yellowing seedling of cotton

A technology of stem tip meristem and etiolated seedlings, applied in the field of plant genetic engineering, can solve the problems of long transformation cycle, difficult plant regeneration, low efficiency, etc.

Inactive Publication Date: 2014-05-28
SHANDONG UNIV
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Problems solved by technology

[0003] Difficulty in plant regeneration, strong genotype dependence, long transformation cycle and low efficiency are the main factors restricting the genetic transformation of cotton.

Method used

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  • Method for improving conversion rate of foreign gene of stem tip meristem of yellowing seedling of cotton
  • Method for improving conversion rate of foreign gene of stem tip meristem of yellowing seedling of cotton
  • Method for improving conversion rate of foreign gene of stem tip meristem of yellowing seedling of cotton

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Embodiment 1

[0039] Embodiment 1, apply the method of the present invention to obtain transgenic cotton

[0040] 1. Obtaining recombinant Agrobacterium

[0041] The target gene TsVP (genbank registration number AY436553) was recombined into the plant binary expression vector pCAMBIA1300-TsVP-als.

[0042]The plasmids used to construct plant expression vectors are: pCAMBIA1300-P35SMCSTnos–als (abbreviation: pCPA) and pCAMBIA1300-Pubi-TsVP-Tnos-als (abbreviation: pCUA-TsVP), in which pCPA contains CaMV35S promoter, herbicide resistance selection marker gene als and a multiple cloning site (MCS), pCUA-TsVP contains the TsVP gene. Digest the pCUA-TsVP plasmid DNA with XbaI, recover the TsVP fragment, connect it to the pCPA multiple cloning site to identify the insertion direction, and obtain the plant expression vector pCAMBIA1300-P35S-TsVP-Tnos–als ( figure 1 ).

[0043] The pCAMBIA1300-P35S-TsVP-Tnos–als vector was transformed into Agrobacterium tumefaciens LBA4404 to obtain recombinant A...

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Abstract

The invention discloses a method for improving conversion rate of a foreign gene of a stem tip meristem of a yellowing seedling of cotton. According to the method, the stem tip meristem of the yellowing seedling of cotton is taken as a receptor, the apical meristem of the stem tip is scratched in a 'cross' manner or in a shape like a Chinese character 'mi' (means rice) with depth of 0.6-1.0mm, and the target gene is introduced into the cotton by a agrobacterium-mediated method, so as to obtain a transgenic plant. The method has advantages of high conversion rate and good repeatability. Experiments prove that the average conversion rate of the cotton converted by the method is improved by 30% in comparison with that of the cotton converted by the unimproved method, so that the method has very important value for genetic improvement of the cotton.

Description

technical field [0001] The invention relates to a method for increasing the conversion rate of exogenous genes, in particular to a method for increasing the conversion rate of exogenous genes in shoot apex meristems of cotton etiolated seedlings, belonging to the field of plant genetic engineering. Background technique [0002] Since Umbeck et al. (Umbeck et al. Bio / Technology, 1987, 5:263-266) successfully obtained transgenic cotton plants through the Agrobacterium-mediated method in 1987, the Agrobacterium-mediated method has gradually become the most common transformation method in the world. One of the means, and has made great progress (Wu et al, 2005, Plant Breeding, 124(2): 142-146; Guo et al. Biologia Plantarum. 2007, 51(2): 242-248). So far, few articles have been published on genetic transformation of cotton. The main reason is that cotton callus is generally difficult to regenerate after transgenic, and the tissue culture time for cotton somatic embryogenesis and...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/82A01H5/00
Inventor 张可炜陈修贵张举仁
Owner SHANDONG UNIV
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