摘要
本研究建立一种Nested PCR技术检测HBV DNA,即采用内外两对引物分别进行连续两次扩增,使检测极限由一次PCR的10^(-2)Pg提高到10^(-5)Pg;经^(32)P标记寡核苷酸探针作Southern转移杂交以及BgIⅡ作酶切分析证实两次扩增均为特异性扩增。通过抗-HBe阳性、单项抗一HBc阳性和单项抗-HBs阳性三种血清的HBV DNA的检测,表明该方法确能检出标准PCR所不能检出的极低水平的HBV感染,对提高乙型肝炎的诊断水平及更准确地评价药物疗效有重要意义。
A nested polymerase chain reaction (PCR) technique for amplification of the hepatitis B virus DNA was established. Two steps of the reaction were included: in the first step the sequences were amplified by the outer primers, and products of the first amplification were reamplified by the nested primers in the second steps. This technique was found to be highly sensitive and capable of detecting HBV DNA up to l0-6 Pg level. The specificity of the amplified bands was confirmed by Southern blot hybridization analysis using a radiolabled oligonucleotide probe and enzyme cutting by Bgl Ⅱ . Anti-HBe positive, anti-HBc positive alone, and anti-HBs positive alone sera were tested by this technique for HBV DNA, the results showed that the extremely low levels of HBV DNA in these sera were detected but couldn't be found, positive by standard PCR.
出处
《中山医科大学学报》
CSCD
1992年第3期48-52,共5页
Academic Journal of Sun Yat-sen University of Medical Sciences
关键词
聚合酶链反应
乙型肝炎病毒
DNA
polymerase chain reaction
hepatitis B virus
DNA
diagnosis
