艾滋病毒核酸探针的制备

Preparation of HIV Nucleic Acid Probe

采用PXY104(含化学合成的HIV-env26肽基因4重串联体结构)为材料。温和裂解法提取质粒DNA,制备电泳纯化,Hind Ⅲ和EcoR Ⅰ酶解,低融点琼脂糖凝胶电泳回收HIV-DNA片段作为核酸探针;用[α-^(32)P]dATP通过缺口移位法标记,比放射性为4.05—6.69×10~7cpm/μgDNA通过分子杂交能测出lPg的靶DNA,初步试验结果表明,在本文试验条件下,可用该探针检测与之互补的核酸分子。

The plasmid pXY104 (containing HIV-env 26 peptide gene, chemicaly synthesized and cloned in our laboratory) was used as the starting material for pre pasing of HIV nucleic acid probe . The plasmid DNA was extracted with mild lysis method and then purified by means of preparative electrophoresis on agarose gel. The purified plasmid DNA was digested by HindⅢ and EcoR Ⅰ . The HIV-DNA fragment was recovered from Low-melting-temperature agarose and labeled with α-32pdATP using nick translation method,The specific radioactivity of the probe is 4.05-6.69×107cpm/μgDNA. Molecular hybridization using this probe showed that lpg of target DNA could be detected.Preliminary results showed that 32P-labeled HIV-DNA is a suitable probe for searching the complementary nucleic acid molecule.