CSFV陕西分离株SXCDK全基因的克隆与分析

Cloning and analysis of the complete genome of CSFV isolate SXCDK from Shaanxi province

【目的】分离猪瘟病毒（CSFV）SXCDK株,测定其全基因组序列,研究其分子特性及与其他毒株的亲缘关系,为猪瘟病毒分子流行病学研究积累材料。【方法】从陕西省西安市采集疑似猪瘟病料进行病毒分离,获得猪瘟病毒SXCDK株。根据GenBank上发表的猪瘟病毒全基因序列及相关参考文献,合成11对引物,应用RT-PCR技术分11段扩增CSFV分离株SXCDK基因,拼接后获得其全基因组（GenBank登录号：GQ923951）。利用DNAstar、ClustalX1.83和Mega 4.1等软件,对分离株SXCDK与其他猪瘟病毒毒株的核苷酸和氨基酸序列进行比较分析。【结果】CSFV陕西分离株SXCDK基因组全长12 296 nt,由373 nt的5′非翻译区、编码3 899个氨基酸的11 697 nt开放阅读框和226 nt的3′非翻译区组成。分离株SXCDK和其他毒株的核苷酸序列同源性为83.6%~96.2%,氨基酸序列的同源性为91.3%~97.8%。从进化的角度来看,分离株SXCDK与台湾分离株96TD的亲缘关系最近,两者之间的进化距离为0.038,分离株SXCDK与2006年陕西省分离株SXYL2006进化距离较远,两者之间的进化距离为0.065。CSFV SXCDK株属于2.1a亚群毒株。【结论】分离株SXCDK是我国内地首株已确定全基因序列的猪瘟病毒2.1a亚群毒株,与同亚群的96TD亲缘关系最近。

【Objective】 The study was to isolate CSFV strain SXCDK,determine the whole genome sequence,study its molecular characteristics and the relationship with other strains,accumulated research data for molecular epidemiology of classical swine fever epidemic strain virus.【Method】 The CSFV strain SXCDK was obtained after virus isolation from a suspected patholgical material of CSF collected from Xi＇an,Shaanxi province.According to the genome sequences of classical swine fever virus（CSFV） published in GenBank and the reference,eleven pairs of specific primers were synthesized.By using RT-PCT,the genome of a CSFV isolate SXCDK was gained after eleven fragments had been successfully amplified（GenBank accession：GQ923951）.Genome comparison of the SXCDK isolate with other known CSFV isolates was performed and analyzed by using software such as DNAstar,ClustalX 1.83 and Mega 4.1.【Result】 The genome of a CSFV isolate SXCDK,isolated from Shaanxi province,is 12 296 nt in length.It is composed of a 373 nt 5′terminal untranslated region（UTR）,a 11 697 nt open reading frame（ORF） encoding a polyprotein of 3 899 amino acids（aa）,and a 226 nt 3′UTR.SXCDK and other reported strains share 83.6%-96.2% identity at the nucleotide level and 91.3%-97.8% identity at the amino acid level.From an evolutionary point of view,isolate SXCDK is closely related to Taiwan isolate 96TD,with a pairwise distance of 0.038,distantly related to the isolate SXYL2006,which is also isolated from Shaanxi province,with pairwise distance 0.065.【Conclusion】 Isolate SXCDK is the first CSFV subgroup 2.1a strain whose genome sequence has been determined in the mainland of China,has the highest genetic relationship with 96TD,which also belongs to this subgroup.