[Prokaryotic expression of Bm86 gene of Boophilus microplus and optimization of the expression condition]

Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi. 2009 Dec;27(6):531-3.
[Article in Chinese]

Abstract

A pair of specific primers was designed based on the reported Bm86 gene of Boophilus microplus,the Bm86 gene was cloned by PCR using the plasmid pMD18-T-Bm86 as templates, and subcloned into the prokaryotic plasmid pGEX-4T-1. The recombined plasmid was transformed into E. coli BL21(DE3) and followed by expression of the protein induced by different concentration of IPTG for different time. SDS-PAGE showed that the recombinant plasmid pGEX-4T-1/Bm86 expressed a fusion protein Bm86-GST (Mr 94 000) after being induced with IPTG. High level expression of Bm86-GST was found at 1 mmol/L IPTG condition a fter incubation for 8 h at 37 degree C, and the expression level of the recombinant Bm86-GST reached up to 29% of total E coli proteins Western-blotting analysis showed that the recombinant Bm86-GST was recognized by the rabbit anti-B. microplus positive serum.

Publication types

  • English Abstract
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blotting, Western
  • Cloning, Molecular
  • Gene Expression
  • Ixodidae / genetics*
  • Membrane Glycoproteins / genetics*
  • Membrane Glycoproteins / immunology
  • Molecular Sequence Data
  • Plasmids
  • Recombinant Proteins / genetics*
  • Recombinant Proteins / immunology
  • Recombinant Proteins / metabolism
  • Vaccines / genetics*
  • Vaccines / immunology

Substances

  • BM86 protein, Boophilus
  • Membrane Glycoproteins
  • Recombinant Proteins
  • Vaccines