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凝胶内贴片电泳:一种用于环境DNA纯化的新方法。

"In-gel patch electrophoresis:" a new method for environmental DNA purification.

作者信息

Roh Changhyun, Villatte Francois, Kim Byung-Gee, Schmid Rolf D

机构信息

School of Chemical Engineering, Institute of Molecular Biology and Genetics, Seoul National University, Seoul, South Korea.

出版信息

Electrophoresis. 2005 Aug;26(16):3055-61. doi: 10.1002/elps.200410328.

Abstract

Most of the microorganism species are largely untapped and could represent an interesting reservoir of genes useful for biotechnological applications. Unfortunately, a major difficulty associated with the methods used to isolate environmental DNA is related to the contamination of the extracted material with humic substances. These polyphenolic compounds inhibit the DNA processing reactions and severely impede cloning procedures. In this work, we describe a rapid, simple, and efficient method for the purification of genomic DNA from environmental samples: we added a chromatography step directly embedded into an agarose gel electrophoresis. This strategy enabled the DNA extraction from various environmental samples and it appeared that the purity grade was compatible with digestion by restriction enzymes and polymerase chain reaction (PCR) amplifications.

摘要

大多数微生物物种在很大程度上尚未得到开发利用,可能是生物技术应用中有趣的基因库。不幸的是,用于分离环境DNA的方法存在一个主要困难,即提取的材料会被腐殖质污染。这些多酚类化合物会抑制DNA处理反应,并严重阻碍克隆程序。在这项工作中,我们描述了一种从环境样品中纯化基因组DNA的快速、简单且高效的方法:我们在琼脂糖凝胶电泳中直接加入了一个层析步骤。这种策略能够从各种环境样品中提取DNA,并且其纯度等级似乎与限制酶消化和聚合酶链反应(PCR)扩增兼容。

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