Abstract:

A pair of primers specific to ChIFN-γ gene were designed and synthesized according to the known cDNA sequences from GenBank,and ChIFN-γ gene was cloned and amplified by reverse transcripition-polymerase chain reaction （RT-PCR） from the total mRNA in the lymphocyte of chicken blood stimulated with ConA for 8 hours. RT- PCR product was cloned into pMD18-T vector. The recombinant plasmid was identified by digestion with restriction endonucleases and the nucleotide sequence was determined. The ChIFN-γ gene was a 495 bp open reading frame encoding a polypeptide of 164 amino acids. The gene was inserted into the expression vector pPICZa-A,which had been cleaved by Kpn I and Not I. The recombinant vector was transfered into yeast Pichia pastoris strain X-33 ,and the ChIFN-γ gene was successfully expressed after induced by methanol. The activity of the recombinant ChIFN-γ was detected by the Western-blotting. The results showed that the recombinant yeast ChIFN-γ was a protein with reactivity.

Key words: chicken y interferon, gene cloning, secretory expression