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Expression, purification and application of E.coli O157:H7 flagellin H7 antigen segment

A flagellin and protein technology, which is applied in the field of genetic engineering technology and detection reagents, can solve the problems of long experiment time and unsuitability for rapid detection, and achieve the effects of easy purification, avoiding interference, good antigenicity and specificity

Active Publication Date: 2014-05-21
中国人民解放军东部战区疾病预防控制中心
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Problems solved by technology

However, these methods more or less have shortcomings such as long experiment time and not suitable for on-site rapid detection.

Method used

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  • Expression, purification and application of E.coli O157:H7 flagellin H7 antigen segment
  • Expression, purification and application of E.coli O157:H7 flagellin H7 antigen segment
  • Expression, purification and application of E.coli O157:H7 flagellin H7 antigen segment

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Embodiment Construction

[0052] Detailed description of the embodiments of the present invention:

[0053] E. coli O157: Analysis, gene synthesis and expression of H7 flagellin H7 epitope

[0054] Computer analysis E. coli O157: The entire amino acid sequence of H7 Flic protein, screened out E. coli O157: The strong epitope in the H7 Flic protein, using the codons preferred by bacteria, chemically synthesize a new gene fragment of the strong epitope of the Flic protein. The gene fragment was cloned into the plasmid pGEX4T-2 Bam HI / Eco The RI site, consistent with the translation framework of the start codon on the vector, can express a fusion protein. Transform the recombinant plasmid E. coli BL21 (DE3), screened and obtained the engineered bacteria that highly express Flic protein, and the expressed Flic protein accounts for about 30% of the total bacterial protein.

[0055] Materials and Methods

[0056] 1. Strains and plasmids: E. coli BL21 (DE3) and expression vector pGEX4T-2 are kept in our labo...

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Abstract

The invention discloses expression, purification and application of (i) E.coli ( / i) O157:H7 flagellin H7 antigen segment, relating to the field of genetic engineering technology, vaccine and diagnostic reagent. According to computer analysis, strong antigen epitope in (i) E.coli ( / i) O157:H7Flic protein is screened out, totally 131 amino acid, from 215th amino acid to 345th amino acid, codon preferred by prokaryote is selected for chemical synthesis of a brand-new gene sequence of the antigen epitope, by genetic engineering technology, the gene segment is expressed, and the strong antigen epitope segment of the (i) E.coli ( / i) O157:H7Flic protein is prepared. The expressed protein can be used for detection of (i) E.coli ( / i) O157:H7 infected antibody and preparation of monoclonal antibody and polyclonal antibody.

Description

Technical field [0001] The present invention involves E. Coli O157: The expression, purification and application of H7 whipped protein H7 antigen fragment, through chemical synthesis E. Coli O157: H7 whipped protein H7 antigen new gene fragment, using genetic engineering technology to prepare recombinant protein.Through computer analysis, the whipped protein H7 antigen fragment containing a strong antigen surface is screened, and a codon that prefers the primary biological preference to chemically synthesize a new gene sequence. Use genetic engineering technology to express E. Coli O157: The detection of H7 antibodies and the preparation of monoclonal antibodies, which are the fields of genetic engineering technology and detection reagents. Background technique [0002] E. Coli O157: H7 is a pathogenicity E. Coli One of the serum types, the infectious diarrhea caused by it is the extremely serious intestinal infectious disease found in recent years.In addition to causing diarrhe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/245C07K16/12C12N15/31C12N15/70G01N33/569
Inventor 李越希李素芹蔡冉李丙军潘英陈乐如周洁许桂丽徐悦玥张素芬马颖袁敬宇
Owner 中国人民解放军东部战区疾病预防控制中心
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