Abstract
Functional characterizations and molecular dissections of long noncoding RNAs (lncRNAs) are critical to understand their involvement in the cellular regulatory network. LncRNAs exert their effects through functional RNA domains that interact with other molecules, including proteins, DNA, and RNA. Here, we describe experimental procedures for generating genomic deletions in a human haploid cell line using the CRISPR/Cas9 system. This method can be applied to examine functions of lncRNAs and their domains by establishing knockout and partial deletion mutant cell lines. In addition, we describe a CRISPR-mediated knockin method for artificial tethering of partner RNA-binding proteins to lncRNAs and its use to validate lncRNA-mediated functions.
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Acknowledgments
This work was supported by grants from the Ministry of Education, Culture, Sports, Science, and Technology of Japan (to T.Y. [26891001, 15K18474, 17K15058, 19H05250, and 19K06479] and T.H. [26113002, 17H03630, 17K19335, 20H00448, 20H05377 and 19K22374]), the Mochida Memorial Foundation for Medical and Pharmaceutical Research (to T.Y.), and Tokyo Biochemical Research Foundation (to T.H.).
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Yamazaki, T., Hirose, T. (2021). CRISPR-Mediated Mutagenesis of Long Noncoding RNAs. In: Cao, H. (eds) Functional Analysis of Long Non-Coding RNAs. Methods in Molecular Biology, vol 2254. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1158-6_18
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DOI: https://doi.org/10.1007/978-1-0716-1158-6_18
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-1157-9
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