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Abstract
RNA localization is an important event that is essential for the polarization and differentiation of a cell. Although several methods are currently used to detect localized RNAs, a simplified detection system has not yet been developed for Schizosaccharomyces pombe. In the present study, we describe a new vector system for the visualization of localized RNAs in S. pombe using a U1A-tag-GFP system. A pREP1-U1A-tag vector plasmid to express U1A-tagged RNA and a pREP2-U1AGFP plasmid to produce a U1A-GFP fusion protein were constructed for this system. Since the U1A-GFP protein binds U1A-tagged RNA, fluorescence is observed at the location of U1A-tagged RNA in cells expressing both of these. The nucleolar localization of U3 snoRNA was successfully detected using this system, and a novel RNA localized at the DNA region of the nucleus was found by screening localized RNAs. This system will accelerate the study of localized RNAs in S. pombe.
Graphical abstract
A, A simplified vector system for visualization of localized RNAs in S. pombe. B, A novel RNA localized in the DNA region of the nucleus, B1199, was found by screening.
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Acknowledgments
We thank all the members of Tani’s laboratory at Kumamoto University for their helpful discussions.
Disclosure statement
No potential conflict of interest was reported by the authors.
Notes
Abbreviations: ONPG, 2-Nitrophenyl-b-D-galactopyranoside; RT-PCR, reverse transcriptase polymerase chain reaction; bp, base pairs.