Kilari S, Rai A
Division of standardization, Indian Veterinary Research Institute, Izatnagar.
Indian J Biochem Biophys. 1993 Jun;30(3):144-50.
Bovine herpesvirus 1 DNA has been isolated by SDS lysis of the virus purified from potassium tartrate (10-50%) density gradient centrifugation. The quality and quantity of viral DNA was checked by UV spectrophotometry and ethidium bromide stained agarose gel electrophoresis. The 0.4 kb Hin dIII'O' fragment of BHV-1 DNA was selectively cloned into Hin dIII cut pUC9 plasmid DNA (2.665 kb). Recombinants were screened by white/blue colonies as well as Hin dIII restriction enzyme analysis. On restriction endonuclease analysis of recombinant plasmid DNA (p-BH-0) with several restriction enzymes, viz., Sau 3A, Hin fI, Rsa I, Sal I, Dra I, Bgl I, Bgl II, Sma I, Hpa I, Stu I, Mlu I, Xho I, Kpn I, Hae III, Eco RI, Bam HI, Pst I, Pal I, revealed insert viral DNA having sites for Hin fI, Hae III, Rsa I, Sma I, only. Further, the partial restriction map of the recombinant plasmid DNA was constructed using above enzymes.
通过对从酒石酸钾（10 - 50%）密度梯度离心纯化的病毒进行SDS裂解，分离出了牛疱疹病毒1型DNA。通过紫外分光光度法和溴化乙锭染色的琼脂糖凝胶电泳检测病毒DNA的质量和数量。将牛疱疹病毒1型DNA的0.4 kb HindIII 'O'片段选择性克隆到经HindIII切割的pUC9质粒DNA（2.665 kb）中。通过白色/蓝色菌落以及HindIII限制性内切酶分析筛选重组体。用几种限制性内切酶，即Sau 3A、HinfI、Rsa I、Sal I、Dra I、Bgl I、Bgl II、Sma I、Hpa I、Stu I、Mlu I、Xho I、Kpn I、Hae III、Eco RI、Bam HI、Pst I、Pal I对重组质粒DNA（p - BH - 0）进行限制性内切酶分析，结果显示插入的病毒DNA仅具有HinfI、Hae III、Rsa I、Sma I的位点。此外，使用上述酶构建了重组质粒DNA的部分限制性图谱。
