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Method for recombination, amalgamation and expression of series antimicrobial peptide gene

A fusion expression, antimicrobial peptide technology, applied in recombinant DNA technology, DNA/RNA fragments, introduction of foreign genetic material using vectors, etc., can solve the problems of low proportion, low expression and low solubility, and achieve high expression The effect of promoting soluble expression and increasing expression

Inactive Publication Date: 2008-12-10
FEED RESEARCH INSTITUTE CHINESE ACADEMY OF AGRICULTURAL SCIENCES
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Problems solved by technology

[0003] The present invention is a method for expressing tandem antimicrobial peptide gene through recombinant fusion, expressing multiple copies of antibacterial peptide gene by fusion with TrxA, not only utilizing multiple copies to increase the ratio of target polypeptide in the expression product, but also increasing the tandem antimicrobial peptide gene through fusion protein The expression of the antimicrobial peptide gene and the promotion of soluble expression overcome the shortcomings of the low proportion of the target polypeptide in the fusion protein in the single-copy antimicrobial peptide gene method of fusion protein expression and the low expression and solubility of the direct tandem method of the target polypeptide gene

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  • Method for recombination, amalgamation and expression of series antimicrobial peptide gene
  • Method for recombination, amalgamation and expression of series antimicrobial peptide gene
  • Method for recombination, amalgamation and expression of series antimicrobial peptide gene

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Embodiment Construction

[0019] Construction of recombinant expression vector and identification of transformants

[0020] The nucleotide sequence of the gene was designed according to the codon preference of Escherichia coli, the sense and antisense strands were synthesized respectively, and a multi-copy gene was formed through the non-mirror symmetry complementary cohesive ends, and the pre-linker containing the BamH I restriction site and the Hind containing Post linker ligation of the III restriction site. The multi-copy gene with the linker was digested with HindIII and BamH I, and then ligated with the linearized pET32a vector to construct the recombinant plasmid expression vector pET32a-(LfcinB 15-W4, 10) n , After transforming Escherichia coli DH5α, positive recombinants containing different copy numbers of coding genes were obtained by colony PCR identification and screening according to the fragment size of the amplified product.

[0021] Figure 1 shows the results of agarose gel electropho...

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Abstract

The invention relates to a method for expressing a cascade antibacterial peptide gene by recombination and fusion. The multi-copy antibacterial peptide gene is expressed through the fusion with thioredoxin TrxA, thereby not only making use of the multiple copies to improve the proportion of a target polypeptide in an expression product, but also improving the expression amount of the cascade antibacterial peptide gene and promoting soluble expression through fusing protein, thereby overcoming the defects of low polypeptide proportion in the fusion protein by using a fusion expression single-copy antibacterial peptide gene method, and low expression amount and low solubility by using a direct cascade target polypeptide gene method. The method can realize the efficient expression of the cascade antibacterial peptide gene. Under optimal conditions, the total expression amount of the recombinant and fused antibacterial peptide exceeds 2mg / mL (extracting solution).

Description

technical field [0001] The technology of the present invention relates to recombinant expression technology, especially the recombinant fusion expression of tandem antimicrobial peptide genes Background technique [0002] At present, the recombinant expression of antimicrobial peptide genes mainly includes the following two ways: one is to express a single copy of antimicrobial peptide genes by fusion with GST or TrxA, which has the advantages of increasing the expression of antimicrobial peptide genes and promoting soluble expression. The proportion of target polypeptides in fusion proteins is low; 2. Directly express by tandem target polypeptide genes. Although the proportion of target polypeptides in the expression products is high, the lack of GST or TrxA fusion proteins can improve the expression of antimicrobial peptide genes. Promote soluble expression and other advantages. SUMMARY OF THE INVENTION [0003] The present invention is a method for recombinant fusion e...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/62C12N15/69
Inventor 王建华田子罡杨雅麟滕达
Owner FEED RESEARCH INSTITUTE CHINESE ACADEMY OF AGRICULTURAL SCIENCES
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