SARS冠状病毒Mc区克隆及序列分析  

Cloning and sequence analysis of SARS-CoV Mc region

作　　者：刘志伟[1] 赵卫[2] 龙北国[2] 

机构地区：[1]南方医科大学南方医院检验科,广州市510515 [2]南方医科大学公共卫生与热带医学学院微生物学系,广州市510515

出　　处：《实用医学杂志》2007年第12期1785-1787,共3页The Journal of Practical Medicine

基　　金：广东省医药科研基金资助项目(编号:A2004378)

摘　　要：目的:构建SARS冠状病毒(SARS-CoV)GD322株M基因膜内区(Mc区)重组表达质粒,并对目的序列进行序列分析。方法:根据GenBank中公布的SARS-CoVTor2株M基因序列,设计一对引物,采用RT-PCR法从SARS-CoVGD322株基因组中扩增Mc区,克隆至pET-32a(+)载体中,转化大肠杆菌BL21后测序,利用ClustalX分析所测序列翻译的氨基酸与81株SARS-CoVMc区翻译的氨基酸序列的差异。结果:测序结果表明该区与Tor2株对应核苷酸序列同源性为100%。与已收集的81株SARS-CoVMc区所译氨基酸(Mc蛋白)相比,与77株(占95.1%)Mc蛋白完全同源,与4株(占4.9%)仅有一个氨基酸改变。结论:本试验成功构建SARS-CoVMc区重组表达质粒;SARS-CoV各毒株间Mc蛋白氨基酸序列差异极小,提示利用任一毒株Mc蛋白制备单抗和疫苗具有普遍意义。To construct a recombinant expressing plasmid of SARS-CoV GD322 Mc region and analyze its sequences. Methods Based on the sequences of Tor2 M gene published on GenBank, we generated a pair of primers, amplified Me fragments from the isolate of SARS-CoV GD322 using RT-PCR, then inserted the Me fragments into expression vetor pET-32a（＋）, and then transformed E. coli BL21 with the chimeric plasmid, finally sequenced the products. The amino acid sequences of Me protein from the SARS-CoV GD322 isolate and those from 81 SARS-CoV isolates were analyzed by Clustal X. Results The sequencing result showed the nueleotide sequence in GD322 had a homology of 100% with the corresponding sequence in Tor2;The amino acid sequence of Mc protein translated by GD322 Mc region was completely homologous to that of 77 out of the 81 SARS-CoV isolates collected from GenBank and only had one different amino acid as compared with the remaining 4 isolates. Conclusions The recombinant expression plasmid of SARS-CoV Mc region is successfully constructed. There is little difference in the amino acid sequence of Mc protein among the isolates, suggesting the monocloned antibody or the vaccine for SARS-CoV generated from the Mc proteins of any isolate has a universal significance.

关 键 词：SARS病毒 Mc蛋白 序列分析 克隆 

分 类 号：R373[医药卫生—病原生物学]