猪繁殖与呼吸综合征病毒 TJM 株5′和3′非翻译区克隆与测序

摘要

In order to obtain the accurate sequences of 5′untranslated region (5′UTR)and 3′untranslated region (3′UTR)of porcine reproductive and respiratory syndrome virus(PRRSV)TJM strain,we used rapid amplification of cDNA ends (RACE )to cloning the sequence.The results showed that the tech-niques of 3′and 5′RACE could amplify the terminal regions of PRRSV effectively,and the 493 bp and 750 bp segments of PRRSV were obtained.Furthermore,sequence analysis compareisons with the wild strain TJ (GenBank Number:EU860248.1)and other strains showed that there are 4 mutations compared with TJ in 5′UTR as same as XJu-1 strain,and the former 3 are identical with some of Asian stains.While the 3′UTR had no change.Moreover,according to predicting the secondary structure of RNA,the mutations of the 5′UTR of TJM strain could change the secondary structure of RNA,and it plays a key role in viral rep-lication and rescued.The accurate sequence of the 5′ and 3′ terminal regions of TJM strain laid a sound foundation for analyzing the function and structure of genome and construction of full-length infectious clone of PRRSV.%为了获得猪繁殖与呼吸综合征病毒（PRRSV）TJM 株5′非翻译区（5′UTR）和3′非翻译区（3′UTR）的准确序列，采用 cDNA 末端快速扩增（RACE）技术进行克隆测序。结果表明，5′和3′末端快速扩增技术（5′RACE 和3′RACE ）能很好地扩增猪繁殖与呼吸综合征病毒的末端序列，分别获得长度为493 bp和750 bp 的片段。与 NCBI 中已发表的原野毒株 TJ 株序列（EU860248．1）及其他毒株比对分析发现，TJM株5′UTR 较 TJ 株有4处突变，突变后与 XJu-1株序列完全一致，其中，前3处突变与许多亚洲毒株相同，3′UTR 序列没有改变。通过预测 RNA 二级结构发现，TJM 株5′UTR 核苷酸序列的改变能够影响 RNA 二级结构，而该二级结构是病毒复制和拯救的关键。TJM 株5′和3′末端序列的获得，为病毒基因组功能结构分析和全长感染性克隆的构建奠定了基础。