猪传染性胃肠炎病毒SC-H株M基因的克隆与序列分析

Cloning and Sequence Analysis of M Gene of Transmissible Gastroenteritis Virus SC-H Strain

参照Genbank已发表的TGEV的核酸序列设计一对扩增M基因的引物,经RT-PCR扩增了SC-H株的M基因cDNA片段,插入pMD18-T构建重组质粒pMD-M,对pMD-M进行了酶切鉴定及核苷酸测序,将M基因核苷酸序列及推导的氨基酸序列与参考毒株进行了比较分析。结果表明SC-H株M基因扩增片段长度为852bp,包含789bp的M基因编码区,编码262个氨基酸;SC-H株与TS、96-1933、FS772/70、HN2002、TFI、Purdue及TO14株的M基因核苷酸序列同源性为97.8%、94.2%、97.2%、97.8%、99.7%、96.8%及98.2%,氨基酸同源性为96.6%、92.0%、95.4%、96.6%、99.2%、95.8%及97.7%,表明不同毒株的M基因高度保守,系统进化分析显示SC-H株与Prudue株有较近的亲缘关系。

A pair of primers for M gene was designed according to the published cDNA sequence of transmissible gastroenteritis virus（TGEV） on Genbank ,which was used to amplify the complete cDNA fragments of M gene by reverse transcription polymerase chain reaction （RT-PCR）. The amplified fragments were cloned into pMD18-T vector and the recombinant plasmid pMD-M was identified by restriction enzyme analysis and sequencing, the nucleotide sequences and deduced amino acid sequences were analyzed in comparison with some other published TGEV strains. The results show the amplified fragments were 852 bp, the M gene was 789 bp and coding 262 amino acids. Compared with the strains Purdue, TO14, TS, HN2002, FS772/70, TFI and 96--1933, the homology of nucleotide sequences was 97.8%, 94.2%, 97.2%, 97.8%, 99.7%, 96.8% and 98.2% respectively, the homology of nucleotide sequences was 96.6%, 92.0%, 95.4%, 96.6%, 99. 2%, 95.8% and 97.7% respectively, which show that M gene of different TGEV strains was highly conservative. The phylogenetic analysis show SC-H strains has a nearest genetic relationship with Strain Prude.