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Nucleic Acids Research Advance Access originally published online on May 30, 2008
Nucleic Acids Research 2008 36(12):e72; doi:10.1093/nar/gkn295
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Nucleic Acids Research, 2008, Vol. 36, No. 12 e72
© 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

Inversing the natural hydrogen bonding rule to selectively amplify GC-rich ADAR-edited RNAs

Rodolphe Suspène1,2, Myrtille Renard1,2, Michel Henry1,2, Denise Guétard1,2, David Puyraimond-Zemmour1,2, Agnès Billecocq3, Michèle Bouloy3, Frederic Tangy2,4, Jean-Pierre Vartanian1,2 and Simon Wain-Hobson1,2,*

1Unité de Rétrovirologie Moléculaire, 2CNRS URA 3015, 3Unité de Génétique Moléculaire des Bunyaviridés and 4Laboratoire de Génomique Virale et Vaccination, Institut Pasteur, 28 rue du Dr Roux, 75724 Paris cedex 15

*To whom correspondence should be addressed. Tel: +33 1 45 68 88 21; Fax: +33 1 45 68 88 74; Email: simon{at}pasteur.fr

Received January 28, 2008. Revised April 25, 2008. Accepted April 29, 2008.

DNA complementarity is expressed by way of three hydrogen bonds for a G:C base pair and two for A:T. As a result, careful control of the denaturation temperature of PCR allows selective amplification of AT-rich alleles. Yet for the same reason, the converse is not possible, selective amplification of GC-rich alleles. Inosine (I) hydrogen bonds to cytosine by two hydrogen bonds while diaminopurine (D) forms three hydrogen bonds with thymine. By substituting dATP by dDTP and dGTP by dITP in a PCR reaction, DNA is obtained in which the natural hydrogen bonding rule is inversed. When PCR is performed at limiting denaturation temperatures, it is possible to recover GC-rich viral genomes and inverted Alu elements embedded in cellular mRNAs resulting from editing by dsRNA dependent host cell adenosine deaminases. The editing of Alu elements in cellular mRNAs was strongly enhanced by type I interferon induction indicating a novel link mRNA metabolism and innate immunity.


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