PRRSV广西分离株GXYL-1403全基因克隆与序列分析  

Cloning and Sequence Analysis Full-length Genome of PRRSV Isolate GXYL1403 in Guangxi

作　　者：林思远[1] 洪绍锋[1] 黄加滨 韦莹[1] 陈樱[1] 黄伟坚[1] 韦祖樟[1] 

机构地区：[1]广西大学动物科学技术学院动物传染病研究室,广西南宁530005

出　　处：《动物医学进展》2016年第2期44-49,共6页Progress In Veterinary Medicine

基　　金：国家自然科学基金项目(31372444);广西大学科研基金项目(XGZ130959)

摘　　要：对本实验室所分离的1株猪繁殖与呼吸综合征病毒（PRRSV）毒株GXYL-1403进行全基因组克隆和序列分析。参考已发表的扩增全长PRRSV序列的13对特异性引物,用RT-PCR方法对病毒基因组进行分段扩增。将扩增得到的PCR片段克隆到pMD18-T载体上,获得的重组质粒经PCR鉴定后进行序列测定。通过软件对所扩增序列进行拼接,获得GXYL-1403的全长序列,并对GXYL-1403进行比较和遗传进化分析。结果表明GXYL1403株基因序列全长为15 018bp,与国内外多株PRRSV毒株进行同源性比较发现,与VR-2332的相似性为89.1%,与TJ、JXA1、HEB1、HUB1、HUB2、TP株同源性达到了97.9%～98.1%。另外,GXYL-1403分离株nsp2蛋白含有高致病性PRRSV特异的1＋29个氨基酸的缺失。特别重要的是,在缺失29个氨基酸区域的上游,含有连续20个氨基酸缺失,在下游出现了连续74个氨基酸的缺失。遗传进化分析发现美洲型PRRSV主要分为3个亚群,GXYL-1403跟高致病性PRRSV毒株属于同一分支,证实了GXYL-1403株为美洲型PRRSV毒株。To clone and analyse the full-length genome of a PRRSV strain GXYL-1403 isolated in our lab, 13 pairs of primers published were used for RT-PCR to amplify the full-length genome of GXYL-1409. The PCR fragments were cloned into pMD18-T vector. The amplicons were identified by emzyme digestion and further sequenced. The full-length genome was obtained through DNA splicing by DNAstar software. The phylogenic tree of GXYL1403 and the reference strains was constructed based on the full-length genome. The results showed that the complete genome of GXYL1403 consisted of 15 018 bp. The complete genome of GXYL-1403 shared 89.1% identity with VR-2332 and shared 97.9%-98.1% identity with HP-PRRSV strains TJ,JXA1, HEB1, HUB1, HUB2, TP. The phylogenic analysis based on the full-length genome re- vealed that PRRSV strains were classified into 3 subgroups and GXYL-1403 was confirmed as HP-PRRSV. It was suggested that GXYL1403 belongs to type II PRRSV. GXYL-1403 strain contained a deletion of 1 ＋ 29 aa in nsp2 coding region. More importantly, GXYL-1403 strain contained a deletion of 20 aa in nsp2 coding region, which was located upstream of 29 aa deletion. Beside, a continuous deletion of 74 aa e- merged following the 29 aa deletion in nsp2 region.

关 键 词：猪繁殖与呼吸综合征病毒 克隆 序列分析 遗传进化分析 

分 类 号：S852.659.6[农业科学—基础兽医学] Q785[农业科学—兽医学]