In vitro production and multiplex quantification of 2',3'-cyclic phosphate-containing 5'-tRNA half molecules.

Methods : a Companion to Methods in Enzymology
Takuya KawamuraYohei Kirino

Abstract

RNA cleavages by many ribonucleases generate RNA molecules that contain a 2',3'-cyclic phosphate (cP) at their 3'-termini, and many cP-containing RNAs (cP-RNAs) are expressed as functional molecules in cells and tissues. 5'-tRNA half molecules are representative examples of functional cP-RNAs, playing important roles in various biological processes. We here show in vitro production of cP-containing 5'-tRNA half molecules that is able to prepare abundant synthetic cP-RNAs enough for functional analyses. Furthermore, we report a multiplex TaqMan RT-qPCR method which can simultaneously quantify multiple cP-containing 5'-tRNA half species. The method enabled us to efficiently quantify 5'-tRNA halves using samples with limited amounts, such as human plasma samples, revealing drastic enhancement of 5'-tRNA half levels at approximately 1,000-fold in patients infected with Mycobacterium tuberculosis. These in vitro production and multiplex quantification methods can be applied to any cP-RNAs, and they provide cost-effective, in-house techniques to accelerate expressional and functional characterizations of 5'-tRNA halves and other cP-RNAs.

References

Nov 11, 1987·Nucleic Acids Research·J F MilliganO C Uhlenbeck
Feb 21, 1998·Nucleic Acids Research·M SprinzlS Steinberg
Jun 13, 2002·Nucleic Acids Research·Heike SchürerMario Mörl
Feb 20, 2004·Nucleic Acids Research·Agnieszka WichlaczJerzy Ciesiołka
Dec 31, 2008·FEBS Letters·Hanjiang FuXiaofei Zheng
Apr 1, 2009·The Journal of Cell Biology·Satoshi YamasakiPaul Anderson
Feb 5, 2010·The Journal of Biological Chemistry·Mohamed M EmaraPaul Anderson
Aug 23, 2011·Molecular Cell·Pavel IvanovPaul Anderson
Mar 20, 2012·Cellular and Molecular Life Sciences : CMLS·Johannes PopowJavier Martinez
Sep 6, 2012·Proceedings of the National Academy of Sciences of the United States of America·Markus EnglertJimin Wang
Jul 27, 2014·The EMBO Journal·Sandra BlancoMichaela Frye
Sep 16, 2014·FEBS Letters·Paul Anderson, Pavel Ivanov
Nov 19, 2014·Proceedings of the National Academy of Sciences of the United States of America·Pavel IvanovPaul Anderson
Jul 1, 2015·Proceedings of the National Academy of Sciences of the United States of America·Shozo HondaYohei Kirino
Jun 25, 2017·Nucleic Acids Research·Shozo HondaYohei Kirino
Mar 20, 2018·Trends in Microbiology·James A TsatsaronisEmmanuelle Charpentier
May 20, 2018·Proceedings of the National Academy of Sciences of the United States of America·Klaas E A MaxThomas Tuschl
Dec 13, 2018·Frontiers in Genetics·Megumi ShigematsuYohei Kirino
Sep 25, 2019·Proceedings of the National Academy of Sciences of the United States of America·John B MoldovanJohn V Moran
May 14, 2020·RNA Biology·Megumi Shigematsu, Yohei Kirino
May 28, 2020·Nature Reviews. Molecular Cell Biology·Killian O'BrienXandra O Breakefield

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