猪嵴病毒441株3D基因的克隆及序列分析

Molecular Cloning and Analysis of the 3'UTR of Porcine Kobuvirus441 by 3'RACE

根据GenBank中登录的猪嵴病毒(porcine kobuvirus,PKV)核苷酸序列设计引物,利用RT-PCR技术及3'RACE法扩增猪嵴病毒441株(swKoV CH441株)3D基因。成功扩增了swKoV CH441株的3D基因,3'非编码区(3'untranslated region,3'UTR)及Poly(A),扩增的基因片段长度分别为1 064、1 046和592 bp,包括3A、3B、3C、3D基因序列,3'UTR序列和一个至少含有29个Poly(A)的尾巴。序列分析结果表明,swKoV CH441株3D基因全长1 407 bp,编码468个氨基酸,分子质量约为53 369.29 D,理论等电点为6.12;3'UTR位于终止密码子TGA之后,长166 bp;swKoV CH441株与其他猪嵴病毒3D基因核苷酸序列相似性在92.2%-93.4%,氨基酸一致性为97.9%-99.4%。遗传进化分析表明,swKoV CH441株与国内株亲缘关系较近,与匈牙利株亲缘关系较远。

The special primers of porcine kobuvirus （ PKV ） were designed according to the sequence logged in GenBank. The 3D gene was amplified by RT-PCR and 3＇ RACE. The sequence analysis showed that the amplified specific sequences were 1 064 bp, 1 046 bp and 592 bp in length respectively, including 3A, 3B, 3C, 3D region, 3＇untranslated region （ 3＇ UTR ） and at least a 29-poly （ A ） tail. The 3D region was 1 407 bp in length, encoding 468 amino acid （ aa ） with a molecular weight of 53 369.29 D, pI of 6.12. The 3＇ UTR was 166 bp in length after the terminator codon TGA. The aligning analysis showed that the 3D gene of swKoV CH441 shared high nncleotide acid and amino acid identity with the other porcine kobuvirus strains sharing 92.2% to 93.4% and 97.9% to 99.4%, respectively, In the phylogenetic tree, swKoV CH441 was closer with Chinese strains but further with Hungarian strains.