聚合酶链反应技术结合酶切杂交研究单项抗乙型肝炎病毒核心抗原阳性者的乙型肝炎病毒感染

以聚合酶链反应（ＰＣＲ）扩增单项抗乙型肝炎病毒核心抗原（ＨＢｃ）阳性血清的乙型肝炎病毒（ＨＢＶ）ＤＮＡ。内外两对引物均选自ａｄｒ、ａｄｗ和ａｙｗ３种亚型ＨＢＶ基因组Ｃ基因区的共有序列，扩增产物分别长４２８ｂＰ和６６４ｂＰ，共含一个ＢｇｌⅡ酶切点，酶切后前者产生２９９ｂＰ和１２５ｂＰ、后者产生４０６ｂｐ和２５４ｂｐ各一个限制性片段。以另一条基因位置处于内引物对之间的寡核苷酸制备３２Ｐ－５＇末端标记的探针对之进行Ｓｏｕｔｈｅｒｎ转移杂交，鉴定了扩增产物的特异性。结果从单项抗ＨＢｃ阳性的８／４２例慢性肝炎和２／１２倒无症状受测者血清中检出ＨＢＶＤＮＡ，提示现症低水平病毒复制和潜在的传染性。

Amplification of Hepatitis B Virus(HBV) DNA by Polymerase Chain Reaction(PCR) in Serum which is Solely Positive to Anti-hepatitis B Core Antigen(HBc)

Five oligonucleohdes nested in the C open reading frame(ORF) of the HBV genome and co-conserved by the subtpe adr, adw and ayw were synthesized 4 of which were pairedly used as primers in the polymerase chain reaction to amplify HBV DNA in serum which is solely positive to anti-HBc. The product of the outer primer pair is 664 bp in length, and of the inner 428 bp, both sharing a Bgl II restriction site in their sequences. After digestion with the enzyme, the 664 bp product yielded a 406 bp and a 254 bp fragment. and the 428 bp product yielded a 299 bp and a 125 bp. All of them were determined by Southern-blot hybridization that useing the other oligonucleotide as a probe, of which the sequence was nested between the inner pair and the 5' terminus labelled with 32P-dATP. Using these techniques. HBV DNA was detected in 8 of 42chronic hepatitis and 2 of 12 healthy carriers, indicating low-level viral replication as potential infectivity.