Cloning And Prokaryotic Expression Of RongChang Porcine Interleukin-6(IL-6)

The cDNA sequence encoding RongChang porcine interleukin-6 (PIL-6) was cloned from the PBMC stimulated with Con A by means of using the specific primers based on the reported sequence in GenBank(AF309651). The cDNA was connected to the PMD18-T plasmid, then proved to be IL-6 with enzyme digestion and sequence identification. The full length of the cloned cDNA , was 741 bp with an ORF composed of 639 nucleosides, which encodes 212 amino acids. It also showed there was high homology between the Rong Chang PIL-6 and other IL-6 sequence published from the GenBank, which was an identity of 99. 8%-100%, and the indentity of deduced amino acids was 99.5%-l00%. Meanwhile,the Hydrophobicity , antigenic index and surface probability analysis of PIL-6 protein were the same as the feature of the PIL-6 protein.By the technology of DNA recombination, the RIL-6 cDNA was cloned into Prokaryotic Expression plasmids PET-32(a)+ vector, and was transformed into Ecoli. The recombinant plasmid was induced to expression through the 1mM concentration of IPTG. The results of SDS-PAGE showed that the expression products were about 44KD and reached their expression peaks about 3 hours after inducing and the expression quatity of fusion protein had 49. 7% ratio of total thalline protein, while the E. coli without inducing with IPTG...