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CRISPR Epigenome Editing in Human Cells using Plasmid DNA Transfection and mRNA Nucleofection Delivery

CRISPR Epigenome Editing in Human Cells using Plasmid DNA Transfection and mRNA Nucleofection Delivery

Published on: May 30, 2025

Screening Regulatory Element Function with CRISPR/Cas9-based Epigenome Editing.

Tyler S Klann^1,2, Gregory E Crawford^2,3, Timothy E Reddy^1,2,4

¹Department of Biomedical Engineering, Duke University, Durham, NC, USA.

Methods in Molecular Biology (Clifton, N.J.)

|March 11, 2018

Summary

This summary is machine-generated.

We developed CRISPR/Cas9-based epigenomic regulatory element screening (CERES) to identify gene regulatory elements controlling traits and diseases. This method enables high-throughput functional annotation of regulatory elements in their native genomic context.

Keywords:

CRISPR/Cas9 Epigenetics Epigenome editing Gene expression profiling High-throughput screening

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Area of Science:

Genomics
Molecular Biology
Epigenetics

Background:

Genomic regulatory elements are crucial for gene expression, influencing traits and diseases.
Identifying and understanding the function of these elements is a significant challenge in current research.

Purpose of the Study:

To develop a novel high-throughput screening method for assessing regulatory element activity.
To enable functional annotation of regulatory elements within their native genomic context.

Main Methods:

CRISPR/Cas9-based epigenomic regulatory element screening (CERES) was developed.
The protocol involves gRNA library design and cloning, construction of endogenous reporter cell lines using CRISPR/Cas9-mediated knock-in of fluorescent proteins, and screen design.
Recovery of the gRNA library for enrichment analysis is a key step.

Main Results:

CERES provides a robust method for screening regulatory element activity.
The protocol facilitates high-throughput functional annotation of putative regulatory elements.
CRISPR/Cas9 technology is leveraged for precise genome engineering.

Conclusions:

The developed CERES protocol addresses the technological need for efficient regulatory element screening.
This method offers a valuable tool for researchers studying gene regulation, traits, and diseases.
The protocol is broadly applicable for implementing genome engineering in functional genomics studies.