A simple and rapid technique of template preparation for PCR

文献类型： 外文期刊

第一作者： Liu, Yunyun

作者： Liu, Yunyun;Chen, Jia;Cheng, Yi;Xu, Jianping;Li, Zhimin;Liu, Yunyun;Chen, Jia;Cheng, Yi;Xu, Jianping;Li, Zhimin;Li, Yi;Li, Xinwen;Zhang, Zhengbing;Xu, Xiumei;Lin, Yufeng;Xu, Jianping

作者机构：

关键词： simple PCR template preparation; microorganism; potassium hydroxide; long fragment amplification; high through-put PCR

期刊名称：FRONTIERS IN MICROBIOLOGY （ 影响因子：6.064； 五年影响因子：6.843 ）

ISSN：

年卷期： 2022 年 13 卷

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收录情况： SCI

摘要： Many techniques have been developed for extracting DNA, but most are often complex, time-consuming, and/or expensive. In this study, we describe a simple, rapid and cost-effective technique for preparing DNA template for PCR. This technique involves 0.1 M potassium hydroxide treatment at 100 & DEG;C for 10 min followed by centrifugation. The suspended centrifuged sediments were shown as excellent templates for PCR. Templates prepared using this technique worked for diverse microorganisms belonging to bacteria, fungi and oomycetes and their amplification efficiencies were comparable to/better than those prepared using common but relatively more complex, time-consuming, and/or expensive methods, including commercial DNA extraction kits. Furthermore, this technology is suitable for high-throughput batch processing and for amplifications of long DNA fragments. Flow cytometry and scanning electronic microscopy analyzes showed that this technique generated primarily damaged cells and cell-bound DNA, not free naked DNA. This technique provides a great convenience for simple PCR template preparation.

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