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Construction method of recombinant expression vector of plant gene

A technology of gene recombination and expression vector, applied in the direction of plant gene improvement, botanical equipment and methods, biochemical equipment and methods, etc., can solve problems such as failure to clone, and achieve the effect of reducing difficulty and strong salt tolerance

Inactive Publication Date: 2004-11-17
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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Problems solved by technology

Prior art The splicing sequence of BADH cDNA (GenBank registration number: AY256971) has been cloned from Abatrifolia trifoliata, but failed to be cloned into a plant expression vector in order to create new salt-tolerant germplasm

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  • Construction method of recombinant expression vector of plant gene
  • Construction method of recombinant expression vector of plant gene

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Embodiment Construction

[0034] 1. Materials and Methods

[0035] 1.1 Materials

[0036] 1.1.1 Plant material

[0037] Atrigus trifoliata is sown in vermiculite and cultured in Hongland nutrient solution. After the seedlings grow 2 true leaves, some seedlings are treated with salt. The initial concentration of treatment is 50mM NaCl, which is increased by 50mM every 3 days until 500mM, and The shoots were grown for one week in a solution of 500 mM NaCl.

[0038] 1.1.2 Strains and plasmids

[0039] pMD18-T Vector is a product of TakaRa Company. PCAMBIA2301 (Hajdukiewicz P, Svab Z, Maliga P (1994) The small, versatile Ppzp family of Agrobacterium binary vectors for plant transformation. Plant Mol. Biol. 25: 989-994) and JM109 (currently available in biochemical companies) are produced by our laboratory save. LBA4404 (currently available at Biochemical Company) was provided by Dr. Wang Huazhong from Nanjing Agricultural University.

[0040] 1.1.3 Main chemical reagents and enzymes

[0041] Trizol ...

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Abstract

The invention relates to a construction method of recombinant expression vector of plant gene, in particular the construction of Atriplex triangularis oxyneurine aldehyde dehydrogenase gene expression vectors, which comprises, utilizing RT-PCR technique to start from halophytic vegetation Atriplex trianglaris total RNA for expanding and cloning BADH gene, connecting the BADH gene to the pCAMBIA2301 polyclonal sites by utilizing BamHI and SacI enzyme cutting sites, and connecting 35S promotor and nos terminator respectively to the upstream and downstream polyclonal sites of the BADH, thus obtaining the expression vector pCBAMATBADH682, which is now used for rape conversion from bacillus through medium guide.

Description

1. Technical field [0001] The invention relates to a method for constructing a plant gene recombination expression vector, which belongs to a method for constructing a plant expression vector of Atrigus trifoliata betaine aldehyde dehydrogenase gene, and is specially used for plant expression using Atrigus trifoliata betaine aldehyde dehydrogenase gene to create new salt-tolerant Germplasm. 2. Technical Background [0002] Atriplex triangularis is a plant of the Chenopodiaceae genus. It is naturally distributed on the edge of coastal swamps. It is an annual leafy plant that can withstand direct watering from seawater. The leaves are edible and have strong salt and drought tolerance. The general response of organisms to abiotic stress is to accumulate low molecular weight organic solutes compatible with cell metabolism, such as sucrose, mannitol, proline, trehalose, and quaternary ammonium compounds. Betaine is a class of quaternary ammonium compounds. [0003] Glycine beta...

Claims

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Application Information

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IPC IPC(8): C12N1/21C12N15/29C12N15/53C12N15/70C12N15/82C12Q1/68
Inventor 何晓兰刘桂华朱卫民张保龙倪万潮钦佩侯喜林
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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