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Nucleic Acids Research Advance Access originally published online on February 19, 2008
Nucleic Acids Research 2008 36(5):e32; doi:10.1093/nar/gkn074
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Nucleic Acids Research, 2008, Vol. 36, No. 5 e32
© 2008 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Methods Online

The use of multiple displacement amplification to amplify complex DNA libraries

Melissa J. Fullwood, Jack J. S. Tan, Patrick W. P. Ng, Kuo Ping Chiu, Jun Liu, Chia Lin Wei and Yijun Ruan*

Genome Institute of Singapore, Agency for Science, Technology and Research (A*STAR), 60 Biopolis Street, Genome #02-01, Singapore 138672

*To whom correspondence should be addressed. Tel: (65) 6478 8073; Fax: (65) 6478 9059; Email: ruanyj{at}gis.a-star.edu.sg

Received December 14, 2007. Revised January 17, 2008. Accepted February 5, 2008.

Complex libraries for genomic DNA and cDNA sequencing analyses are typically amplified using bacterial propagation. To reduce biases, large numbers of colonies are plated and scraped from solid-surface agar. This process is time consuming, tedious and limits scaling up. At the same time, multiple displacement amplification (MDA) has been recently developed as a method for in vitro amplification of DNA. However, MDA has no selection function for the removal of ligation multimers. We developed a novel method of briefly introducing ligation reactions into bacteria to select single insert DNA clones followed by MDA to amplify. We applied these methods to a Gene Identification Signatures with Paired-End diTags (GIS-PET) library, which is a complex transcriptome library created by pairing short tags from the 5' and 3' ends of cDNA fragments together, and demonstrated that this selection and amplification strategy is unbiased and efficient.

The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors.


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