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Nat Methods. 2015 Sep;12(9):835-837. doi: 10.1038/nmeth.3478. Epub 2015 Jul 27.

Efficient and quantitative high-throughput tRNA sequencing.

Author information

1
Department of Biochemistry & Molecular Biology, the University of Chicago, Chicago, IL 60637, USA.
2
Institute for Cellular and Molecular Biology, the University of Texas at Austin, Austin, TX 78712, USA.
3
Department of Chemistry, the University of Chicago, Chicago, IL 60637, USA.
4
Institute of Biophysical Dynamics, the University of Chicago, Chicago, IL 60637, USA.
5
Howard Hughes Medical Institute, the University of Chicago, Chicago, Illinois 60637, USA.
#
Contributed equally

Abstract

Despite its biological importance, tRNA has not been adequately sequenced by standard methods because of its abundant post-transcriptional modifications and stable structure, which interfere with cDNA synthesis. We achieved efficient and quantitative tRNA sequencing in HEK293T cells by using engineered demethylases to remove base methylations and a highly processive thermostable group II intron reverse transcriptase to overcome these obstacles. Our method, DM-tRNA-seq, should be applicable to investigations of tRNA in all organisms.

PMID:
26214130
PMCID:
PMC4624326
DOI:
10.1038/nmeth.3478
[Indexed for MEDLINE]
Free PMC Article
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