Li Guangxing, Zeng Yan, Yin Jiechao, Lillehoj Hyun S, Ren Xiaofeng
College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
Hybridoma (Larchmt). 2010 Feb;29(1):1-6. doi: 10.1089/hyb.2009.0053.
The full-length chicken interferon-gamma (CHIFN-gamma) gene was amplified by reverse transcription-PCR using total RNA extracted from the spleen cells of white Leghorn chicken, a local Chinese breeding species. A truncated CHIFN-gamma gene without the N-terminal signal peptide sequence was cloned into prokaryotic expression vector pET30a, resulting in a recombinant plasmid pET-30a-CHIFN-gamma. After the recombinant plasmid was transformed into host cells BL21(DE3)pLysS, the expression of CHIFN-gamma was induced by isopropyl beta-D-thiogalactoside (IPTG). Rabbit antiserum was raised using the soluble CHIFN-gamma as immunogen. Immunoreactivities of the CHIFN-gamma and its antiserum were investigated using immunoblotting and ELISA. Moreover, the antiviral effect of the CHIFN-gamma was analyzed. Our data indicate that the CHIFN-gamma is biologically active.
利用从中国地方培育品种白来航鸡脾脏细胞中提取的总RNA，通过逆转录聚合酶链反应（RT-PCR）扩增出全长鸡γ干扰素（CHIFN-γ）基因。将不含N端信号肽序列的截短型CHIFN-γ基因克隆到原核表达载体pET30a中，得到重组质粒pET-30a-CHIFN-γ。将重组质粒转化到宿主细胞BL21(DE3)pLysS中后，用异丙基-β-D-硫代半乳糖苷（IPTG）诱导CHIFN-γ表达。以可溶性CHIFN-γ作为免疫原制备兔抗血清。利用免疫印迹法和酶联免疫吸附测定法（ELISA）研究CHIFN-γ及其抗血清的免疫反应性。此外，还分析了CHIFN-γ的抗病毒作用。我们的数据表明CHIFN-γ具有生物活性。
