Abstract
CRISPR-Cas systems of prokaryotes are immune defense mechanisms against foreign nucleic acids via RNA-guided endonuclease activities. Small regulatory RNAs (sRNA) are essential for controlling various aspects in gene expression and function in bacteria. Here, we describe a protocol that utilizes T4 RNA ligase 1 (single-stranded RNA ligase) to link two base-paired RNA molecules to investigate potential interaction between sRNA and CRISPR-Cas system. Our goal is to provide readers a detailed method of identifying candidate sRNAs that may regulate CRISPR-Cas adaptation and/or other functions through unbiased screening and validation.
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Acknowledgments
This work was supported by National Institutes of Health Grants R01 AI109317-01A1, R01 AI138203-03, P20 GM113123, and P20 GM103442; this work was also supported by UND Post-Doc Pilot Grant and The American Association of Immunologists through a Careers in Immunology Fellowship; this work is also supported by National Nature Science Foundation of China Grants 32000033, 81530063, and 82020108021.
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Lin, P., Pu, Q., Wu, M. (2021). An Approach to Proximity Ligation by T4 RNA Ligase to Screen sRNA That Regulate CRISPR-Cas Systems. In: Islam, M.T., Molla, K.A. (eds) CRISPR-Cas Methods. Springer Protocols Handbooks. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1657-4_19
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DOI: https://doi.org/10.1007/978-1-0716-1657-4_19
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Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-1656-7
Online ISBN: 978-1-0716-1657-4
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