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Improvement of expression quantity of foreign proteins by fusion label

A technology that fuses tags and proteins, and is applied in the introduction of foreign genetic material using vectors, microorganism-based methods, microorganisms, etc., and can solve the problems of low expression and failure to meet the requirements of industrial production.

Inactive Publication Date: 2013-04-10
JIANGNAN UNIV
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  • Abstract
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AI Technical Summary

Problems solved by technology

However, for some exogenous proteins, the expression level is still very low or cannot meet the requirements of industrial production despite the above optimization

Method used

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  • Improvement of expression quantity of foreign proteins by fusion label
  • Improvement of expression quantity of foreign proteins by fusion label
  • Improvement of expression quantity of foreign proteins by fusion label

Examples

Experimental program
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Embodiment Construction

[0016] 1. Obtaining the first 50 amino acid coding sequences of Bacillus subtilis flagellin Hag

[0017] The hag gene of B. subtilis 168 (Genbank sequence number: M26948) is 915 bp in full length, and is one of the most expressed proteins in Bacillus subtilis. Genomic DNA was extracted from the cultured B. subtilis168 bacterial liquid according to the instructions of the Bacterial Genome Rapid Extraction Kit. PCR primers hag-F and hag50-R were designed according to the hag gene sequence in the whole genome of B.subtilis168 published by NCBI, and NdeI and EcoRI restriction sites were introduced into the upstream and downstream primers respectively. The primers were provided by Sangon Bioengineering (Shanghai) Limited Company Synthesis. Using the extracted genomic DNA as a template, the first 50 amino acid coding sequences of flagellin Hag were amplified by PCR. The amplified product was detected by agarose gel electrophoresis, and the size was consistent with the expectation. ...

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Abstract

The invention belongs to the field of molecular biology, and relates to application of previous 50 amino acids of Bacillus subtilis l68 flagellin Hag, and a novel method for improving the expression quantity of foreign proteins in Bacillus subtilis, and in particular relates to a method for improving the expression quantity of foreign proteins by way of fusion expression. A fusion label Hag 50 coding sequence is fused and recombined with that of the foreign proteins to construct a recombinant plasmid in fusion expression and convert the Bacillus subtilis. The obtained recombinant bacteria have already improved the expression quantity of foreign proteins successfully.

Description

technical field [0001] The invention relates to a new method for increasing the expression of exogenous protein in Bacillus subtilis. More specifically it involves the first 50 amino acids of the Bacillus subtilis flagellin Hag (Hag50) as a fusion tag to achieve overexpression of foreign proteins. Background technique [0002] Escherichia coli has always been the preferred expression system for expressing foreign proteins. However, because foreign proteins are easily degraded by host cell proteases or form inclusion bodies during the expression process, and E. coli has pyrogenic lipopolysaccharide, its application is limited. Due to The genetic background of Bacillus subtilis is clear, and it is a food-grade microorganism. Therefore, in recent years, Bacillus subtilis has developed rapidly as a host for exogenous gene expression and has shown good industrial application prospects. Although Bacillus subtilis is mainly used to achieve secretory expression of exogenous protein...

Claims

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Application Information

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IPC IPC(8): C12N15/75C12R1/125
Inventor 陈卫王光强宋元达陈海琴
Owner JIANGNAN UNIV
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