HBx-DNA探针的研制及在肝癌发生机制研究中的应用  

HBx DNA PROBE PREPARATION AND ITS APPLICATION IN THE STUDY OF HEPATOCARCINOGENESIS

作　　者：高丰光[1] 孙汶生[1] 曹英林[1] 张利宁[1] 宋静[1] 李华芬[1] 阎世坤[1] 

机构地区：[1]山东医科大学微生物学教研室

出　　处：《山东医科大学学报》1997年第1期11-14,共4页Acta Academiae Medicinae Shandong

基　　金：山东省科委科研基金

摘　　要：将非放射性标记物地高辛甙元用随机引物法由ｐＢＲ３２２－２ＨＢＶ／ＥｃｏＲⅠ酶切片段（３．２ｋｂ）制备ＨＢＶ－ＤＮＡ探针及由ＨＢＶ／ＢａｍＨⅠ、ＢｇｌⅡ酶切片段（０．５９ｋｂ）制备ＨＢｘ－ＤＮＡ探针。将Ｄｉｇ－ＨＢＶＤＮＡ探针用斑点杂交、Ｓｏｕｔｈｅｒｎ转膜杂交检测筛选肝细胞性肝癌ＨＢＶ－ＤＮＡ整合型标本，进而用Ｄｉｇ－ＨＢｘＤＮＡ探针检测ＨＢＶ－ＤＮＡ纯整合型肝细胞性肝癌中ＨＢｘ的存在情况。结果显示，７５％（３３／４４）的肝癌ＤＮＡ中有ＨＢＶ－ＤＮＡ存在，其中，以纯整合型方式存在的为６３．６％（２１／３３），混合型为３６．４％（１２／３３），无游离型。纯整合型ＨＢＶ－ＤＮＡ片段中，ＨＢｘ－ＤＮＡ阳性率为９０．５％（１９／２１）。提示ＨＢＶ在肝细胞性肝癌的发生中起着重要的作用，ＨＢＶ－ＤＮＡ可能以片段性整合的方式存在于肝癌细胞染色体上，其中以ＨＢｘ基因的整合为主。HBV 3.2kb fragment was received by digesting recombinated plasmid pBR 322 2HBV with EcoRI,so was HBx 0.59kb fragment by digesting HBV DNA with BamHⅠ,BglⅡ.These fragments were labelled by random primer method with digoxigenin to get HBV DNA probe and HBx DNA probe.HBV DNA was detected in hepatocellular carcinoma(HCC) tissues by dot blot and Southern blot hybridization with the HBV DNA probe.So the positive specimens in which HBV DNA were integrated only were selected.HBx DNA was subsequently detected in the selected specimens with the HBx DNA probe.The results showed:(1) HBV DNA was detected in 75% HCC specimens,and among these specimens integrated type,both integrated and free type were 63.6%(21/33),36.4%(12/33)respectively.There is no free type.(2)HBx DNA was detected in 90.5% HCC specimens of integrated type.The results demonstrated that hepatocarcinogenesis had high relationship with HBV DNA integration,and HBV DNA mainly integrated into chromosome DNA with incomplete virus DNA fragments among which HBx fragment was the major one.

关 键 词：乙型肝炎病毒 肝癌 核酸探针 核酸杂交 

分 类 号：R735.704[医药卫生—肿瘤] R730.231.3[医药卫生—临床医学]