UNIT 15.11 Helicase-Dependent Amplification of Nucleic Acids
Published Online: 11 OCT 2013
Copyright © 2001 John Wiley & Sons, Inc. All rights reserved.
Lab Protocol Title
Current Protocols in Molecular Biology
How to Cite
Cao, Y., Kim, H.-J., Li, Y., Kong, H. and Lemieux, B. 2013. Helicase-Dependent Amplification of Nucleic Acids. Current Protocols in Molecular Biology. 104:15.11:15.11.1–15.11.12.
- Published Online: 11 OCT 2013
Helicase-dependent amplification (HDA) is a novel method for the isothermal in vitro amplification of nucleic acids. The HDA reaction selectively amplifies a target sequence by extension of two oligonucleotide primers. Unlike the polymerase chain reaction (PCR), HDA uses a helicase enzyme to separate the deoxyribonucleic acid (DNA) strands, rather than heat denaturation. This allows DNA amplification without the need for thermal cycling. The helicase used in HDA is a helicase super family II protein obtained from a thermophilic organism, Thermoanaerobacter tengcongensis (TteUvrD). This thermostable helicase is capable of unwinding blunt-end nucleic acid substrates at elevated temperatures (60° to 65°C). The HDA reaction can also be coupled with reverse transcription for ribonucleic acid (RNA) amplification. The products of this reaction can be detected during the reaction using fluorescent probes when incubations are conducted in a fluorimeter. Alternatively, products can be detected after amplification using a disposable amplicon containment device that contains an embedded lateral flow strip. Curr. Protoc. Mol. Biol. 104:15.11.1-15.11.12. © 2013 by John Wiley & Sons, Inc.
- isothermal amplification;
- nucleic acid quantitation;
- lateral flow strip