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A simple and rapid technique of template preparation for PCR

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°C for 1...

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Autores principales: Liu, Yunyun, Chen, Jia, Cheng, Yi, Li, Yi, Li, Xinwen, Zhang, Zhengbing, Xu, Xiumei, Lin, Yufeng, Xu, Jianping, Li, Zhimin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9686307/
https://www.ncbi.nlm.nih.gov/pubmed/36439815
http://dx.doi.org/10.3389/fmicb.2022.1024827
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author Liu, Yunyun
Chen, Jia
Cheng, Yi
Li, Yi
Li, Xinwen
Zhang, Zhengbing
Xu, Xiumei
Lin, Yufeng
Xu, Jianping
Li, Zhimin
author_facet Liu, Yunyun
Chen, Jia
Cheng, Yi
Li, Yi
Li, Xinwen
Zhang, Zhengbing
Xu, Xiumei
Lin, Yufeng
Xu, Jianping
Li, Zhimin
author_sort Liu, Yunyun
collection PubMed
description 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°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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spelling pubmed-96863072022-11-25 A simple and rapid technique of template preparation for PCR Liu, Yunyun Chen, Jia Cheng, Yi Li, Yi Li, Xinwen Zhang, Zhengbing Xu, Xiumei Lin, Yufeng Xu, Jianping Li, Zhimin Front Microbiol Microbiology 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°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. Frontiers Media S.A. 2022-11-10 /pmc/articles/PMC9686307/ /pubmed/36439815 http://dx.doi.org/10.3389/fmicb.2022.1024827 Text en Copyright © 2022 Liu, Chen, Cheng, Li, Li, Zhang, Xu, Lin, Xu and Li. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Microbiology
Liu, Yunyun
Chen, Jia
Cheng, Yi
Li, Yi
Li, Xinwen
Zhang, Zhengbing
Xu, Xiumei
Lin, Yufeng
Xu, Jianping
Li, Zhimin
A simple and rapid technique of template preparation for PCR
title A simple and rapid technique of template preparation for PCR
title_full A simple and rapid technique of template preparation for PCR
title_fullStr A simple and rapid technique of template preparation for PCR
title_full_unstemmed A simple and rapid technique of template preparation for PCR
title_short A simple and rapid technique of template preparation for PCR
title_sort simple and rapid technique of template preparation for pcr
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9686307/
https://www.ncbi.nlm.nih.gov/pubmed/36439815
http://dx.doi.org/10.3389/fmicb.2022.1024827
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