An allylic/acyclic adenosine nucleoside triphosphate for termination of DNA synthesis by DNA template-dependent polymerases.

An allylic adenosine triphosphate analog (AATP) was tested as a substrate for commercially available DNA polymerases. All but one of the enzymes assayed incorporated AATP opposite thymidine (T) with concomitant termination of the elongation reaction. A concentration of only 1 microM was sufficient for complete termination of the polymerization reaction for a short template mediated by Ampli Taq DNA polymerase FS (Taq FS). This result suggests that AATP could be used as a 2',3'-dideoxyadenosine-5'-triphosphate (ddA) surrogate. Kinetics of incorporation revealed that AATP was 48 times less efficiently incorporated than ddA. Furthermore, AATP was used in dye-primer sequencing as a substitute for ddA.本研究将烯丙基腺苷三磷酸类似物（allylic adenosine triphosphate analog, AATP）作为底物，对市售DNA聚合酶开展底物活性测试。在所测试的全部酶中，仅有一种未能将AATP掺入至胸腺嘧啶（thymidine, T）对应的模板位点，且该过程伴随聚合延伸反应的终止。仅需1 μM的AATP浓度，即可使Ampli Taq DNA聚合酶FS（Taq FS）介导的短模板聚合反应完全终止。该结果表明，AATP可作为2',3'-双脱氧腺苷-5'-三磷酸（2',3'-dideoxyadenosine-5'-triphosphate, ddA）的替代底物。掺入动力学实验结果显示，AATP的掺入效率较ddA低48倍。此外，AATP可作为ddA的替代物应用于染料引物测序实验。