Nucleic Acids Research Advance Access originally published online on September 18, 2007
Nucleic Acids Research 2007 35(18):6322-6329; doi:10.1093/nar/gkm657
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Nucleic Acids Research, 2007, Vol. 35, No. 18 6322-6329
© 2007 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.
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Identification of recognition residues for ligation-based detection and quantitation of pseudouridine and N6-methyladenosine
1Department of Biochemistry and Molecular Biology, 2Howard Hughes Medical Institute, University of Chicago, Chicago, IL 60637, 3Department of Chemistry and 4Department of Biochemistry and Biophysics, University of Rochester Medical Center, Rochester, NY 14642, USA
*To whom correspondence should be addressed. Tel: +1 773 702 9312; Fax: +1 773 702 0271; Email: jpicciri{at}uchicago.edu Correspondence may also be addressed to Tao Pan. Tel: +1 773 702 4179; Fax: +1 773 702 0439; E-mail: taopan{at}uchicago.edu
Received June 18, 2007. Revised August 7, 2007. Accepted August 8, 2007.
Over 100 chemical types of RNA modifications have been identified in thousands of sites in all three domains of life. Recent data suggest that modifications function synergistically to mediate biological function, and that cells may coordinately modulate modification levels for regulatory purposes. However, this area of RNA biology remains largely unexplored due to the lack of robust, high-throughput methods to quantify the extent of modification at specific sites. Recently, we developed a facile enzymatic ligation-based method for detection and quantitation of methylated 2'-hydroxyl groups within RNA. Here we exploit the principles of molecular recognition and nucleic acid chemistry to establish the experimental parameters for ligation-based detection and quantitation of pseudouridine (
) and N6-methyladenosine (m6A), two abundant modifications in eukaryotic rRNA/tRNA and mRNA, respectively. Detection of pseudouridylation at several sites in the large subunit rRNA derived from yeast demonstrates the feasibility of the approach for analysis of pseudouridylation in biological RNA samples.
The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors