猪轮状病毒中国分离株JL94 VP7基因的克隆与序列分析

Cloning and sequence analysis of gene encoding outer capsid protein VP7 of porcine rotavirus(RV) JL94 strain isolated in China

用 MA10 4细胞培养增殖了轮状病毒地方分离株 JL94。利用一对根据 Genebank中已发表的轮状病毒 VP7基因c DNA序列而设计并合成的引物 ,通过反转录—聚合酶链反应 (RT- PCR)从 JL94毒株扩增出 VP7基因全长 c DNA。将其插入p MD18- T载体中 ,构建了重组质粒 p MD18- T- JL 94 / VP7,并对其进行了 Hind ,Hind 和 Bam H 的单、双酶切初步鉴定及其核苷酸序列测定 ,证明克隆的 p MD18- T- JL94 / VP7为轮状病毒的 VP7基因。通过核苷酸序列比较 ,JL94 / VP7与 A群猪轮状病毒 C134/ VP7、OSU / VP7、ICB2 185 / VP7、YM/ VP7核苷酸序列同源性分别为 87%、99%、74 %和 83%。

The native JL94 strain of porcine rotavirus was propagated and harvested on MA104 cell. Using a pair of primer designed according to the published cDNA sequence of RV's VP7 gene , the full length gene encoding outer capsid protein VP7 was amplified from JL94 strain by reverse transcription-polymerase chain reaction (RT-PCR). The cDNA of VP7gene was inserted into pMD18-T vector and was identified by sequencing, single and double digestion using HindⅢ, HindⅢ and BamHⅠ.The result of its homology comparison with C134/VP7, OSU/VP7, ICB2185/VP7 and YM/VP7 is 87%, 99%, 74% and 83% respectively.