Journal
NUCLEIC ACIDS RESEARCH
Volume 44, Issue 3, Pages -Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkv971
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Funding
- Combined Funding Application (University at Albany-RNA Institute Capital Project) from the State of New York [X765]
- National Institute on Drug Abuse, National Institute of Health [1R41DA038983-01]
- Research Foundation for SUNY at The University at Albany [1R41DA038983-01]
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Exploration of the epitranscriptome requires the development of highly sensitive and accurate technologies in order to elucidate the contributions of the more than 100 RNA modifications to cell processes. A highly sensitive and accurate ultra-high performance liquid chromatography-tandem mass spectrometry method was developed to simultaneously detect and quantify 28 modified and four major nucleosides in less than 20 min. Absolute concentrations were calculated using extinction coefficients of each of the RNA modifications studied. A comprehensive RNA modifications database of UV profiles and extinction coefficient is reported within a 2.3-5.2 % relative standard deviation. Excellent linearity was observed 0.99227-0.99999 and limit of detection values ranged from63.75 attomoles to 1.21 femtomoles. The analytical performance was evaluated by analyzing RNA modifications from 100 ng of RNA from human pluripotent stem cell-derived neural cells. Modifications were detected at concentrations four orders of magnitude lower than the corresponding parental nucleosides, and as low as 23.01 femtograms, 64.09 attomoles. Direct and global quantitative analysis of RNA modifications are among the advantages of this new approach.
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