鸡γ-干扰素基因的克隆及原核表达  

Cloning and Expression of Chicken IFN-γ Gene

作　　者：芦芝川[1,2] 史耀旭 杨孝朴[1] 

机构地区：[1]甘肃农业大学动物医学院,甘肃兰州730070 [2]中农威特生物科技股份有限公司,甘肃兰州730046

出　　处：《中国畜牧兽医》2012年第3期47-50,共4页China Animal Husbandry & Veterinary Medicine

摘　　要：参照Digby发表的鸡γ-干扰素的核苷酸序列,设计1对引物,应用反转录聚合酶链式反应(RT-PCR)技术,从Con A诱导过的鸡脾淋巴细胞中特异性的扩增出大小约为500bp的基因片段。将该扩增基因进行克隆测序,结果表明其序列与Digby报道的CHIFN-γ基因序列的同源性达100%。将该基因插入原核表达载体pGEX-4T-1中构建成重组表达载体pGEX-CHIFN-γ,并将其导入大肠杆菌BL21中,诱导表达的重组蛋白经SDS-PAGE分析分子质量约为43ku,表明所克隆的CHIFN-γ基因在原核细胞中得到表达。The total RNA was abstracted from the concanavalin A（Con A）-activated spleen lymphocytes,a pair of specific primers was designed by Oligo software based on the sequence of chicken interferon-gamma submitted by digby in GenBank（accession No.U27465）.A 500 bp DNA fragment was amplified by reverse transcription-polymerase chain reaction（RT-PCR） and then ligated into T-easy vector for sequencing.The results showed that the fragment contained the complete open reading frame（ORF） of CHIFN-γ gene.In comparison with GenBank data,the homology of the nucleotide sequence was 100%.The sequence was inserted into pGEX-4T-1 and the recombined expressing vector was constructed,then transformed into Escherichia coli BL21,which were further induced by IPTG and cultured,and a fusion protein was obtained with about 43 ku of molecular weight in SDS-PAGE.This results showed that the cloned CHIFN-γ gene was expressed in prokaryotic cells.

关 键 词：Γ-干扰素 鸡 克隆 表达 

分 类 号：Q78[生物学—分子生物学]