德国牧羊犬γ干扰素基因克隆表达与抗病毒活性测定

Molecular cloning,expression and antiviral activity analysis of CaIFN-γ gene of German shepherd dog

利用RT-PCR扩增德国牧羊犬γ干扰素(CaIFN-γ)的成熟肽。序列分析表明,CaIFN-γ的成熟肽cDNA由432个核苷酸组成,共编码143个氨基酸。构建载体pGEXT-6p-1/CaIFN-γ表达重组CaIFN-γ,经Glutathione Sepharose 4B亲和层析柱纯化后,得到近100%纯度的重组CaIFN-γ,其在犬肾细胞(MDCK)/猪水泡性口炎病毒(VSV)细胞系上的活性可达1.33×106U/mg。

Using the total mRNA from the lymphocytes of German shepherd dog peripheral blood as the template, canine interferon-γ （CaIFN-γ） gene was cloned and amplified by RT-PCR. Sequence analysis showed that the CaIFN-γ gene was composed of 432 nucleotides encoding a mature protein of 143 amino acids and a stop codon. Highly expressed and purified recombinant protein was harvested. Antiviral assays showed that the antiviral activity was 1.33×10^6 U/mg in MDCK/VSV cell lines.