Sequence Analysis and Expression of Membrane Protein Genes of Canine Coronavirus Epidemic Strains

Membrane (M) protein genes of Canine coronavirus (CCoV) domestic strain DXMV, V1 and V2 were successfully amplified, sequenced and analyzed. The full length of all the M gene of DXMV, V1 and V2 strains was 792 bp, encoding 263 amino acids. The initial 17 amino acids as appeared to be a signal peptide. Compared with the strain Insavc-1, the identities of nucleotide sequences and deduced amino acid sequences were 92.6% and 92.5% for strain DXMV, 90.9% and 92.0% for strain V1, 91.6% and 92.3% for strain V2, respectively. The variation was mainly in the one third 5’ regions of M gene, particularly in the regions of the 74-76, 120-124 and 131-135 nucleotides. The identities of nucleotide sequences and deduced amino acid sequences among 5 domestic strains were 96.6% and 96.4%, respectively. Phylogenetic tree based on the M genes showed that most domestic strains belong to CCoV genotype II, while Fox3-1and Rac2-1 are classified into genotype I. The M gene of strain DXMV was subcloned into pET28a expression vector. The recombinant M protein was successfully expressed in BL21(DE3), which amounted to 10.2 percent of the total bacterial protein.