Abstract
Genomic regulatory elements that control gene expression play an important role in many traits and diseases. Identifying the regulatory elements associated with each gene or phenotype and understanding the function of that element remain a significant challenge. To address this technological need, we developed CRISPR/Cas9-based epigenomic regulatory element screening (CERES) for improved high-throughput screening of regulatory element activity in the native genomic context. This protocol includes detailed instructions for design and cloning of gRNA libraries, construction of endogenous reporter cell lines via CRISPR/Cas9-mediated knock-in of fluorescent proteins, overall screen design, and recovery of the gRNA library for enrichment analysis. This protocol will be generally useful for implementing genome engineering technologies for high-throughput functional annotation of putative regulatory elements in their native chromosomal context.
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Acknowledgments
This work was supported by the Thorek Memorial Foundation a US National Institutes of Health (NIH) grants to G.E.C., T.E.R., and C.A.G. (R01DA036865 and U01HG007900), a NIH Director’s New Innovator Award (DP2OD008586) and National Science Foundation (NSF) Faculty Early Career Development (CAREER) Award (CBET-1151035) to C.A.G., and NIH grant P30AR066527. T.S.K. was supported by a NIH Biotechnology Training Grant (T32GM008555).
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Klann, T.S., Crawford, G.E., Reddy, T.E., Gersbach, C.A. (2018). Screening Regulatory Element Function with CRISPR/Cas9-based Epigenome Editing. In: Jeltsch, A., Rots, M. (eds) Epigenome Editing. Methods in Molecular Biology, vol 1767. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7774-1_25
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DOI: https://doi.org/10.1007/978-1-4939-7774-1_25
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