绵羊肺腺瘤病毒内蒙古分离株前病毒全基因组克隆及序列分析

Cloning and Sequencing Analyses of the Complete Genome of the Provirus of the Inner-Mongolia Pandemic Strain of the Jaagsiekte Sheep Retrovirus

为了探究绵羊肺腺瘤病毒(Jaagsiekte sheep retrovirus,JSRV)内蒙古流行株与国际各代表株间的亲缘关系,本研究以内蒙古地区绵羊肺腺瘤病自然病例的肺组织总DNA为模板,克隆gag、pro与pol基因,并应用双酶切的方法将其与本实验室先期制备并保存的LTR、env基因连接起来,得到了含有JSRV内蒙古分离株前病毒全基因组重组质粒pMD-JSRV。序列分析结果表明,JSRV内蒙古分离株前病毒全基因组全长7 690bp,具外源性JSRV典型的分子特征:1在gag基因中含Sca I酶切位点;2核衣壳蛋白区有2个保守的可形成锌指结构的"CCHC"基序;3env基因编码的TM区胞浆尾部包含保守的特异性"YXXM"基序。将其进行同源性分析,结果表明该株病毒属JSRV-II型,与分离自美国的代表株AF105220间亲缘关系较近,同源性达95%。本研究为进一步探讨JSRV内蒙古分离株基因组结构特点与其致病机制间的关系奠定了基础。

To investigate the kinship between the Inner Mongolia pandemic strain and representative strains of the Jaagsiekte sheep retrovirus （JSRV）, total DNA from the lung tissue of a JSRV-infected sheep in In- ner Mongolia was used to clone fragments of gag, pro and po! genes. The recombinant plasmid pMD-JS- RV （including complete genomic sequence of the JSRV strain isolated from Inner Mongolia） was construc- ted by linking all the cloned fragments with long terminal repeat （LTR） and env gene fragments （cloned previous and reserved by our research team）. Sequence analyses revealed that the genome was 7690 bp in length and contained several typical molecular markers for exogenous form of JSRV. These included the Sca I restriction site in the gag gene, two predicted ＂CCHC＂ motifs of zinc finger in the encoded nucleo- capsid protein and the predicted ＂YXXM＂ motif in the TM region of Env. Homology analyses showed that the virus strain belonged to the JSRV type II. pMD-JSRV and AF105220 strains shared a nucleotide identi- fication of 95 %. The full length genomic clone of JSRV could provide a molecular basis for an infectious JSRV molecular clone as well as an experimental platform to study the detection and pathogenesis of JS- RV.