Cloning And Prokaryocyte Expression Of Goose Interferon Gene

Interferon can interfere virus in synthesis and reproduction. It is the first defending system of antiviral infection. The study of mammalian interferon already completed, the study of bird interferon also get on quickly, especially in chicken and duck. The study of goose interferon were not reported in any publication.According to the complete nucleotide sequence of duck interferon- α (DuIFN-α )and duck interferon- γ (DuIFN- γ ) published in GenBank, a pair of primers were designed with Oligo 4.1. By reverse transcription-polymerase chain reaction (RT-PCR), a DNA fragment of about 600bp (IFN-α ) and 500bp (IFN- γ) was amplified. And then, these two fragment was cloned into pMD18-T vector. The result of sequencing shows that the open reading frame (ORF) of goose interferon- α (GoIFN-α ) gene was consist of 576bp, and encodes a protein of 191 amino acids. The 30 hydrophobic amino acids at the N-terminus may serve as signal peptide. Mature polypeptide of GoIFN-α seems to consist of 161 amino acids. Sequence comparison with the published DuIFN- α gene in GenBank revealed 96.70% identity at the nucleotide level. Identity at the amino acid level was 93.72%.The open reading frame (ORF) of goose interferon- γ (GoIFN- γ ) gene was consist of 495bp, and encodes a protein of 164 amino acids. The 19 hydrophobic amino acids at the N-terminus may serve as signal peptide. Mature polypeptide of GoIFN- γ seems to consist of 145 amino acids. Sequence comparison with the published DuIFN- Y gene in GenBank revealed 94.95% identity at the nucleotide level. Identity at the amino acid level was 93.29%.But it only shared 31.26% identity in nucleotide level and 6.22% identity in amino acid level between GoIFN- α and GoIFN- y gene. By comparing of the homology with Chicken, Canine and human IFN(IFN- α and IFN- γ) gene in nucleotide respectively and amino acid respectively, identities were very low.The two GoIFN gene was cloned specially into pGEX-6p-1 vector, the recombinant plasmid was transformed into BL21(DE₃)PlysS, and was induced with 1.0mmol/L IPTG for gene expression. The expressed product was identified by SDS-PAGE, a fusion protein about 44ku and 40ku as we expected was found. Then, the GoIFN-γ gene was cloned specially into pET-30a vector between the EcoR I and BamH I cleavage site, the recombinant plasmid was transformed into BL21(DE₃)PlysS, and was induced with 1.0mmol/L IPTG for GoIFN-γ gene expression. The expressed product was identified by SDS-PAGE, a fusion protein about 18ku as we expected was found.This assay result establish the base on further biological study of goose interferon, It also pioneer a new field for study the bird interferon.