Screening Regulatory Element Function with CRISPR/Cas9-based Epigenome Editing.
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.
Duke Scholars
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Related Subject Headings
- Regulatory Sequences, Nucleic Acid
- Red Fluorescent Protein
- RNA, Guide, CRISPR-Cas Systems
- Luminescent Proteins
- K562 Cells
- Humans
- High-Throughput Screening Assays
- HEK293 Cells
- Gene Library
- Gene Knock-In Techniques
Citation
Published In
DOI
EISSN
Publication Date
Volume
Start / End Page
Location
Related Subject Headings
- Regulatory Sequences, Nucleic Acid
- Red Fluorescent Protein
- RNA, Guide, CRISPR-Cas Systems
- Luminescent Proteins
- K562 Cells
- Humans
- High-Throughput Screening Assays
- HEK293 Cells
- Gene Library
- Gene Knock-In Techniques