Terminal restriction fragment length polymorphism monitoring of genes amplified directly from bacterial communities in soils and sediments

被引：12

作者：

Bruce K.D. ^{[1
,2
]}

Hughes M.R. ^{[2
,3
]}

机构：

[1] Division of Life Sciences, Franklin-Wilkins Building, King's College, SE1 8WA London

[2] Department of Biological Sciences, Donnan Labs., University of Liverpool

[3] MWG-BIOTECH UK Ltd., Waterside House, MK6 3BY Milton Keynes, Peartree Bridge

来源：

Molecular Biotechnology | 2000年 / 16卷 / 3期

基金：

英国自然环境研究理事会;

关键词：

Electrophoresis; PCR; Resolution; Sequence variants; Terminal Restriction Fragment Length Polymorphism Profiling;

D O I：

10.1385/MB:16:3:261

中图分类号：

学科分类号：

摘要：

Terminal Restriction Fragment Length Polymorphism (T-RFLP) or Fluorescent Polymerase Chain Reaction/Restriction Fragment Length Polymorphism (FluRFLP) have made a significant impact on the way in which PCR products amplified from mixed community DNA extracts have been assessed. Technically, these approaches are essentially the same. PCR products are generated that contain at one 5′ end label, typically a fluorescent moiety, that will be detected by a DNA sequencing machine. Upon digestion using a specific restriction endonuclease, labeled and unlabeled fragments are generated. This restriction endonuclease is chosen such that following this digestion, each labeled fragment corresponds to a different sequence variant. During electrophoretic separation, the DNA sequencing machine detects only these labeled fragments and therefore detects only the sequence variants. The aim of this article is to describe the protocols and demonstrate that this profiling can be performed using different DNA sequencing machines. The analysis and applications of this approach are also discussed.

引用

页码：261 / 269

页数：8

共 45 条

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