二株广西猪繁殖与呼吸综合征流行毒M基因的克隆和序列分析

Cloning and sequence analysis of M gene of two PRRSV strains epidemic in Guangxi

广西一些养殖场频繁发生以母猪流产、死胎、木乃伊等为特征的流行性传染病,怀疑为猪繁殖与呼吸综合征病毒(PRRSV)所致。利用针对PRRSV的N基因的特异性引物P1和P2,通过RT＿PCR技术对分别从广西贵港市(GXGG)和南宁市(GXNN)收集到的可疑病料进行检测,结果两份为阳性。合成针对M基因的引物M1和M2,分别扩增了GXGG和GXNN两株PRRSV的M基因,并进行克隆和测序,得到长582个核苷酸的目的基因片段。应用DNAStar序列分析软件对所测的两个广西毒株与国内外已发表的ATCCVR＿2332、LV和CH＿la毒株进行同源性比较。分析表明,GXGG与ATCCVR＿2332、LV、CH＿la的核苷酸同源性分别为95 6%、69 5%、97 7%;GXNN与ATCCVR＿2332、LV、CH＿la的核酸同源性分别为94 9%、69 5%、97 3%。对推定的氨基酸序列进行了比较,GXGG与ATCCVR＿2332、LV、CH＿la的氨基酸同源性分别为97 7%、80 5%、97 7%;GXNN与ATCCVR＿2332、LV、CH＿la的氨基酸同源性分别为98 3%、79 9%、97 1%。说明广西流行毒株与ATCCVR＿2332和CH＿la的同源性很高,而与LV毒株的同源性很低。从本研究构建的系统发育树分析,广西流行的PRRSV与ATCCVR＿2332株的亲缘关系比较密切。

In recent, some pig farms in Guangxi occur frequently an infectious disease,which characters are abortion,death of embryo and mummified fetuses.Pathogen of the disease is considered as porcine reproductive and respiratory syndrome virus(PRRSV).With a specific N gene primers of PRRSV,we identified that the samples collected from these famers in Nanning and Guigang were positive by RT-PCR Based on it,we designed two primers M1 and M2.According to the sequence of M gene.The fragments had been successfully amplified from RNA of PRRSV from GXGG and GXNN samples.The amplified fragments were cloned and sequenced.The length of M gene fragments is 582 base pair.Comparison of the homology among GXGG,GXNN,ATCC VR-2332,Lelystad virus and CH-la strain by DNAStar software,the results showed that the homology of nucleotide siquences of nucleotide sequences of GXGG and ATCC VR-2332,Lelystad virus,CH-la were 95.6 %,69.5 %,97.7 %,respectively.and the homology of nucleotide sequences or GXNN and ATCC VR-2332,Lelystad virus,CH-la were 94.9 %,69.5 %,97.3 %,respectively.Comparison of the deduced amino acid sequences showed that strains GXGG and ATCC VR-2331,Lelystad Virus,CH-la were 97.7 %,80.5 %,97.7 %,respectively.Those analysis showed that the homologies among Guangxi,ATCC VR-2332 and CH-la strain were very high,but very low between Guangxi strain and ATCC VR-2332,CH-la or Lelystad virus.And the phylogenetic tree revealed that Guangxi strains perhaps have a close relation to ATCC VR-2332 and CH-la.