Cloning of ORF1 gene of porcine circovirus type 1 and its expression in Escherichia coli

A pair of primers was designed according to the porcine circovirus type 1(PCV1) sequences in GenBank to amplify the open reading frame 1(ORF1) gene of PCV1.Then,the amplicon was cloned into pMD18-T vector to construct recombinant plasmid pMD-ORF1.The identity of nucleotide sequence was 99.6% between the amplified ORF1 gene and that of Guangdong isolate DQ659154,and the identity of the deduced amino acid sequence was 99.4% correspondingly.The ORF1 gene was subcloned into a prokaryo-tic expression vector pET-41a and the recombinant was designated pET-ORF1.The results of SDS-PAGE and Western-blotting indicated that the ORF1 gene was expressed from pET-ORF1 induced by IPTG at high level,and the expressed fusion protein was 67.6 ku approximately in molecular mass.