摘要
建立了生物素标记探针技术，以便用于戊型肝炎病毒（ｈｅｐａｔｉｔｉｓＥｖｉｒｕｓ，ＨＥＶ）的诊断及与肠道病毒的鉴别。用Ａ５４９细胞培养ＨＥＶ８７Ａ株，收集上清，浓缩沉淀，酚／氯仿抽提ＲＮＡ，ＲＴ－ＰＣＫ扩增ｃＣＮＡ，将其直接点于硝酸纤维素膜或扩增产物经琼脂糖凝胶电泳分离后，转移到硝酸纤维素膜上，制备待检样品。以光敏生物素标记ＨＥＶＥＴ１．１４１８６ｐ探针，对硝酸纤维素膜进行点杂交和Ｓｏｕｔｈｅｒｎ印迹杂交。结果表明，制备的ＨＥＶ探针，特别是合成的寡核苷酸ＨＥＶ探针和肠道病毒探针，能够分别特异地与８７Ａ株病毒和肠道病毒Ｅｃｈｏ－１３型病毒的核酸杂交。该方法特异性强，较单独用ＰＣＲ扩增、经琼脂糖凝胶电泳敏感性高，特异性强，且结果能长期保存，可用于戊型肝炎的临床诊断。
Hepatitis E virus(HEV) strain 87A and enterovirus(ETV)echovirus type 13 were inocu-lated onto A549 cell line.Supernatants were collected and concentrated.After extraction of RNAs,the cDNAs were synthesized and amplified by means of PCR(RT-PCR).With photosensitivebiotin lablled probes,HEV derived from clone ET1.1(418 bp)and ETV derived from Echo-13(500bp),Dot and Southern blotting were carried out on the NC membranes.The results showed thatthe DNAs of HEV strain 87A and ETV Echo-13 could be hybridized only with their own specialprobes respectively. There is no homogeneity between the two viruses.The evidence indicates fur-ther that the strain 87A was the pathogen of HEV epidemic in Xinjiang, China.
出处
《军事医学科学院院刊》
CSCD
北大核心
1995年第3期161-166,共6页
Bulletin of the Academy of Military Medical Sciences
关键词
戊型肝炎病毒
聚合酶链反应
鉴别诊断
核酸杂交
hepatitis E virus, polymerase chain reaction
diagnosis, differential, nucleic acidhybridization
electrophoresis,agar gel
