山羊痘病毒基因组DNA的提取及酶切分析

Extraction and Restriction Endonuclease Analysis of Goatpox virus Genomic DNA

对山羊痘病毒贵州分离株(LD株和QL株)和参考株(Y株和B株),采用Vero细胞增殖和差速离心浓缩后,以SDS-蛋白酶K法提取病毒基因组DNA,以3种核酸内切酶EcoRⅠ、HindⅢ和PstⅠ进行酶切图谱分析。接种山羊痘病毒48 h^60 h后,Vero细胞呈现典型的细胞病变;病毒基因组提取样本的纯度在1.8~1.9之间,经琼脂糖凝胶电泳均出现一条纯净的DNA条带;经3种内切酶酶切后,贵州分离株的酶切图谱与参考疫苗毒株Y株基本一致,比参考弱毒株B株少3条~4条酶切片段。这为山羊痘病毒毒力基因的进一步研究奠定了基础。

Goat poxvirus （GPV） LD and QL strains which were isolated from Guizhou province and standard strain Y and B proliferated by cell culture and concentrated by ultracentrifugation. High quality genomic DNA were extracted from goat poxvirus with the SDS-Protease K. Restriction endonuclease maps of 4 goat poxvirus strains were compared by using different restriction endonucleases such as EcoR Ⅰ , HindⅢ, Pst Ⅰ . Vero cells showed the typical cytopathic effect （CPE） 48-60 hours after inoculation with goat poxvirus. The virus genome samples in the purity between 1.8 to 1.9. The agarose gel electrophoresis showed a pure DNA segment. The maps of LD and QL genomic DNA digested by restriction endonucleases EcoR Ⅰ , HindⅢ, Pst Ⅰ showed no differences with Y strains but less 3-4 digested fragments compared with B strain. The results will lay a good foundation for the study of goat poxvirus virulence genes.