Amplification sequencing of gene encoding the protein of bovine interferon-γ

Based on the published nucleotide sequence of bovine interferonγ gene, a pair of RTPCR primers were designed and synthesized. Totle RNA, isolated from peripheral blood T cells of bovine, was used as template to generate complementary DNA by reverse transcription. The 500 bp DNA fragment were amplified by polymerase chain reaction. Sequencing results demonstrated that interferonγgene contains 517 bp with a 501 bp open reading fragment(ORF) encoding a peptide of 166 amino acide whose molecular is 19.4 kd, which is completely accord with reference array. The cloning gene fragment was inserted into expression vector PBV220 when they are cut by BamHI / EcoRI. The recombinant plasmid was transformed into DH5α strain by CaCl2. The SDSPAGE result showed that cloned recombinant protein was expressed in the supernatant with molecular weight of approximated 19.4Ku. The whole positive result has been certified by immunefluorescence technique.