Precision length determination and in silico simulation in PCR of microsatellite repeat sequences
Precision length determination and in silico simulation in PCR of microsatellite repeat sequences
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DOI：
10.1002/elps.202100021
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发表时间：
2021-03
期刊：
影响因子：
2.9
通讯作者：
S. Shioi;Akiyoshi Shimamoto;Yuki Nakagami;Lexin Qin;M. Shimokawa;S. Oda
中科院分区：
文献类型：
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作者：
S. Shioi;Akiyoshi Shimamoto;Yuki Nakagami;Lexin Qin;M. Shimokawa;S. Oda
聚合酶链反应（PCR）尽管很常见，但仍有许多未知的方面。一个例子是微卫星PCR，它现在被广泛用于从生态学到癌症医学的各种目的。由于这类重复DNA序列不仅在体内而且在体外诱导聚合酶滑移，因此微卫星PCR产物包含具有不同长度的DNA片段的复杂组合，因此已被经验性地解释。了解微卫星PCR的主要障碍是固有的不准确性，在毛细管电泳（CE）中的DNA片段的大小，然而，这已经克服了阐明内在的大小误差在每个片段长度范围。其次，首先使用引物延伸测定详细阐明了热稳定聚合酶的滑移性质。此外，使用所获得的滑动参数和我们的原始程序，我们首次在计算机上重建微卫星PCR。因此，复杂微卫星PCR的整个过程已被更清楚地理解。
Despite being commonplace, polymerase chain reactions (PCRs) still contain many unknown aspects. One example is microsatellite PCR, which is now widely used for various purposes from ecology to cancer medicine. Since this category of repetitive DNA sequences induces polymerase slippage not only in vivo but also in vitro, microsatellite PCR products comprise a complex combination of DNA fragments with various lengths and have, therefore, been empirically interpreted. The primary obstacle for understanding microsatellite PCR was the intrinsic inaccuracy in sizing of DNA fragments in capillary electrophoresis (CE), which, however, has been overcome by elucidating intrinsic sizing errors in each fragment length range. Secondly, the slippage properties of the thermostable polymerases were first clarified in detail using primer extension assays. Furthermore, using the obtained slippage parameters and our original program, we have first reconstructed microsatellite PCR in silico. The entire processes of complex microsatellite PCR have, thus, been more clearly understood.