半随机单引物PCR扩增产物的特异性研究

Study on the specificity of amplification products by single-primer PCR

以Ｍ１３ｍｐ１９为模板，寡核苷酸“１２２４”为引物，进行半随机单引物ＰＣＲ扩增；分析其扩增产物的限制性内切酶酶切图谱，推定它们分别在Ｍ１３ｍｐ１９序列中的确切位置；证实引物“１２２４”的３端与模板Ｍ１３ｍｐ１９负链的互补结合，至少需有连续的４个核苷酸，才能产生单引物ＰＣＲ扩增的模板分子，进行有效的ＰＣＲ扩增，并由此决定了扩增产物中各ＤＮＡ片段的大小及其在模板ＤＮＡ序列中的特定位置．

The template M,13mp19 was amplified by the semi-random single-primer PCR with the oligonucleotide primer '1224', The DNA fragments in the amplification products were purified and used as the templates. The latter was then amplified by the same single-primer PCR under the normal condition. The exact regions where these products were located and the starting sites of the specific PCR amplification were determined. It was shown that more than 4 nucleotides at the 3,termination of the primer'1224' integrating complementarily with the template DNA were needed for the production of the template molecules of single-primer PCR and the effective PCR amplification. Moreover, the sites of integration determined the lengths of DNA fragments in the amplification products and their specific locations in the template DNA.