High Level Expression of Grass Carp Reovirus VP7 Protein in Prokaryotic Cells
- Author:
Lanlan ZHANG
;
Jinyu SHEN
;
Chengfeng LEI
;
Xiaoming LI
;
Qin FANG
- Publication Type:Journal Article
- Keywords:
Grass carp reovirus (GCRV);
VP7 protein;
Prokaryotic expression
- From:
Virologica Sinica
2008;23(1):51-56
- CountryChina
- Language:Chinese
-
Abstract:
Sequences analysis revealed Grass carp reovirus (GCRV) s10 was 909 nucleotides coding a 34 kDa protein denoted as VP7, which was determined to be a viral outer capsid protein (OCP). To obtain expressed OCP in vitro, a full length VP7 gene was produced by RT-PCR amplification, and the amplified fragment was cloned into T7 promoted prokaryotic expression vector pRSET. The recombinant plasmid,which was named as pR/GCRV-VP7,was then transformed into E.coli BL21 host cells. The data indicated that the expressed recombinant was in frame with the N-terminal fusion peptide. The over-expressed fusion protein was produced by inducing with IPTG, and its molecular weight was about 37kDa, which was consistent with its predicted size. In addition, the fusion protein was produced in the form of the inclusion body with their yield remaining steady at more than 60% of total bacterial protein. Moreover,the expressed protein was able to bind immunologically to anti-his-tag monoclonal antibody (mouse) and anti-GCRV serum (rabbit). This work provides a research basis for further structure and function studies of GCRV during entry into cells.
