Cloning and Sequence Analysis of Feline Interleukin-18 Gene and Its Expression in Escherichia coli

According to feline interleukin-18 (IL-18) gene sequence reported in GenBank, feline IL-18 gene was amplified by RT-PCR from the feline peripheral blood mononuclear cells (PBMCf) stimulated by concanavalin (ConA). The PCR product was purified and ligatured with pMD18-T. The recombinant plasmid was used for sequencing. The full length of feline IL-18 gene was 579 bp, which encoded 192 amino acids(Genbank accession No.DQ100372). There were no signal peptide and N-glycosylation sites, while had 4 conservative Cys amino acid in the deduced amino acid sequence. Compared with IL-18 genes of other species, the nucleotide sequence of feline IL-18 gene shares 89.8%、88.6%、88.4% and 88.1% identities with IL-18 genes of canine, ovine, bovine and porcine respectively, but displays significant differences with mouse and chicken IL-18 gene sequence which published in GenBank. Meanwhile, the feline IL-18 gene was subcloned into the prokaryotic expressing vector pET28a and recombinant vector was further transformed into Escherichia coli strain BL21(DE3) for expression under the induction of IPTG. The results of SDS-PAGE revealed that it had a molecular weight of 27.5 kD, which amounts to 13.6% in the total protein of the induced bacteria by the assaying of gel scanning.