Sequence Of ORF Gene Of One Prevalent Field Virus Strain Of CSFV (BJCY/96) And Expression Of The Subsection Of E0 Gene Of Classical Swine Fever Virus

The 11 694bp ORF gene of one strain(BJCY/96) of prevalent field virus were amplified by RT-PCR and. Their nucleotide sequences were determined and the amino acid sequences were deduced. Compared with the corresponding region of C, Alfort HCLV and SM strain of CSFV, the nucleotide sequence homologies were 95.8%,97.9%,95.7% and 99.7% respectively, the deduced amino acid consistencies were 97.4%, 98.7%, 97.5% and 99.7% respectively. The mutation rate between the BJCY/96 strain and the HCLV strain were very high, their amino acid mutation rate of the structural protein E0,E1 and E2 were 4.8% ,4.1% and 5.39% respectively. This paper showed accord with the opinion that some strain of prevalent field CSFV have the genetic variation from the strain of HCLV .The three subsection of E0 gene of CSFV of HCLV strain, Shimen strain were amplified by reverse transcription (RT) . The amplified E0 fragments of the two strains were 198bp,243 bp,321 bp respectively in length. And they were cloned into pGEM-T Easy vector. The recombinant plasmids and expression vector pGEX-6P-1 were digested by the same restriction endonucleases. These genes were ligated and transformed into Escherichia coli. The insert position, the size and the reading frame all were corrected by PCR, restriction digestion and the sequence analysis. The result showed that the prokaryotic expression vectors were constructed successfully. Then the recombinants were transformed into BL21 (DE3) for fragments of E0 expression with IPTG inducing. The expressed proteins were measured by SDS-PAGE and western-blotting. The results showed that the three subsection of E0 genes can express successfully in E.coli. The western-blotting results indicated that the expressed protein be recognized by the CSFV positive serum.