直接法提取血清DNA后套式聚合酶链反应扩增乙肝病毒DNA

Direct Extraction of Serum DNA and Amplification of HBV DNA by Nested Polymerase Chain Reaction

为进一步提高ＨＢＶＤＮＡ检测水平，改良了直接法提取血清ＤＮＡ技术，并用套式聚合酶链反应扩增ＨＢＶＤＮＡ。即以裂解液直接提取血清ＨＢＶＤＮＡ，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１０－２ｐｇ提高到１０－６ｐｇ，整个过程３ｈ内完成。经ＢｇｌⅡ酶切分析证实两次扩增均为特异性扩增。

An improved serum DNA extraction method for raising the detection level of HBV DNA was used. HBV DNA form serum could be extracted directly with splitting solution. Two stage amplification could be conducted with outer and inner primers.This method was capable of detecting HBV DNA up to 10 -6 pg level while standard PCR methed could only reach to 10 -2 pg level.The whole process course was completed in three hours. The specificity of amplification was confirmed by Bgl Ⅱ cutting products.This technique can be used in clinical detection and also in studying HBV infection.