Prokaryotic Expression of Bm86 Gene of Boophilus microplus and Optimization of the Expression Condition
- VernacularTitle:微小牛蜱Bm86基因的原核表达与表达条件的优化
- Author:
Miling MA
;
Guiquan GUAN
;
Youquan LI
;
Aihong LIU
;
Qiaoyun REN
;
Qingli NIU
;
Hong YIN
;
Jianxun LUO
- Publication Type:Journal Article
- Keywords:
Boophilus microplus;
Bm86 gene;
Expression
- From:
Chinese Journal of Parasitology and Parasitic Diseases
1997;0(06):-
- CountryChina
- Language: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 expre-ssion of Bm86-GST was found at 1 mmol/L IPTG condition after incubation for 8 h at 37 ℃,and the expression level of the recom-binant 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.
