鸡传染性支气管炎病毒H52毒株S基因的克隆及序列分析  

Cloning and sequencing of the S gene of H52 strain from avian infectious bronchitis virus.

作　　者：程丽琴[1] 周继勇[1] 沈行燕[1] 陈吉刚[1] 陈声明[2] 

机构地区：[1]浙江大学动物预防医学研究所,浙江杭州310029 [2]浙江大学生物科学系,浙江杭州310029

出　　处：《浙江大学学报（农业与生命科学版）》2002年第3期303-306,共4页Journal of Zhejiang University:Agriculture and Life Sciences

基　　金：国家自然科学基金资助项目 (39970 0 30 )

摘　　要：将含有鸡传染性支气管炎病毒 (IBV) H5 2毒株的尿囊液浓缩 ,用 TRIzol试剂抽提病毒 RNA作为反转录 -聚合酶链反应 (RT- PCR)的模板 ,参照已发表的 IBV- Beaudette株 S基因序列 ,设计并合成一对特异性引物 ,扩增得到长度约 3.6 kb的 S基因片段 ,利用引物设计中 Eco R I和 Bam H I酶切位点插入到克隆载体 p Blue Script SK的多克隆位点中 ,转化感受态 E.coli DH5 α。经酶切分析和 PCR等方法鉴定 ,筛选出阳性克隆 ,进行核苷酸序列测定 ,证实获得了 S基因的全长序列。利用 DNAstar等分析软件与 Gen Bank中其它 IBV毒株的核苷酸及其推导的氨基酸进行序列同源性比较分析 ,结果表明 ,H5 2毒株与已报道的核苷酸的同源性在 83.4 5 %～ 99.71%之间 ,氨基酸的同源性在 82 .10 %～ 99.2 6 %之间。Virion RNA from H52 strain of avian Infectious Bronchitis were extracted from the allantoic fluid of inoculated embryonated eggs and used as template for cDNA synthesis by reverse transcription with 3' primer. Polymerase chain reaction was performed with two primers which span the S gene, and an expected product of 3.6 kb was obtained. The PCR product was ligated to pBlueScript SK vector, and its nucleotide sequence was determined by the dideoxy mediated chain termination method. Comparison and analysis of the nucleotide and deduced amino acid sequence of S gene with that of other IBV strains from GenBank was performed by computer with DNAstar and PCgene software. The results showed that the homology of S gene between H52 and other strains was 83.45% 99.71% on nucleotide level, while 82.10% 99.26% identity was found on amino acid level. Phylogenetic tree of the nucleotide sequence of IBV strain S gene was constructed by computer.

关 键 词：鸡传染性支气管炎病毒H52毒株 S基因 克隆 序列分析 

分 类 号：S858.315.3[农业科学—临床兽医学] S852.659.6[农业科学—兽医学]