乙型肝炎病毒的亚克隆及其应用

Subcloning of HBV and the Application on Nucleic Acid Hybridization

目的 应用酶切pBP32 2 HBV ,回收HBVDNA ,将其克隆到高拷贝载体 ,获得亚克隆株 pUC19 HBV ,再酶切、HBVDNA标记探针 ,核酸杂交检测临床血清标本 72份 ,与定量PCR ,定性PCR方法对比。方法 由此获得了亚克隆株pUC19 HBV ,用地高辛标记的HBVDNA探针杂交、定量PCR、定性PCR方法同时检测临床血清标本 72份。结果 阳性标本份数分别为 40、5 1、49份 ,阳性率分别为 5 5 6 %、70 8%、6 8 1%。阳性符合率分别为 88 9%、84 6 %、80 8%。结论 3种HBV核酸检测方法的比较 ,以定量PCR法最为敏感 。

Objective Subcloning strain with whole gene of HBV is applied for nucleic acid hybridization diagnosis and the studies of transgenic tomatoes.Methods In this experiment,clone the HBV DNA of pBR 322 HBV into higher copies vector pUC 19 to get the new strain of pUC 19 HBV.HBV DNA probe labeled with Dicogingenin was made by digesting the pUC 19 HBV with endonuclease and purificafion of HBV DNA.72 cases of patient serum were paralleled detected by nucleic acid hybridization,QPCR and PCR electrophoresis.gain the subcloning strain of pUC 19 HBV,use the DIG HBV probe labeled with Digoxigenin,QPCR and PCR to detect 72 cases serums simultaneously.Results The positive cases were 40,51,49 respectively.The positive percentage was 55 6%,70 8%,68 1% respectively.Conclusions Among the three methods of detecting HBV,QPCR is the most sensitive method,even though the hybridizations is the most specific.