Cloning of Structural Protein VPI Gene of Foot and Mouth Disease Virus and its Expression in Escherichia coli

In this study, the viral RNA was extracted from swine FMDV and then the fragment of VP1 was amplified with a primer pair by RT-PCR. The interest fragmen was inserted into pGEM-Teasy vector. There combinant plasmid was identified by restriction analysis and PCR. It was proved by DNA sequencing that the acquired recombinant contains complete VP1 gene. The homologie of the nucleotide sequence of VP1 gene were 95.9 and 96.2%, respectively comparing with that of strai O/JPN/00 and O-Tibet-99. Afterwards, the complete VP1 gene from the identified recombin an was amplifie with another primer pair containing BamHI and XhoI sites by PCR and digested it with BamHI and XhoI. The expression vector pET28a were digested by BamHI and XhoI, respectively. The target gene VP1 was subcloned into vector pET28a. Positive clones named as pET28a-VP1 with interest gene were identificated by restriction analysis, PCR and DNA sequencing. Then there combinant was transformed into Escherichia coli BL21 (DE3) for VP1 expression. The interest gene was induced to express in E. coli with IPTG. The bacteria containing pET28a-VP1 were collected at different time and subsequently were examined by SDS-PAGE and Western-blotting.