Format

Send to

Choose Destination
Methods Mol Biol. 2019;1935:203-221. doi: 10.1007/978-1-4939-9057-3_14.

Experimental and Computational Approaches for Single-Cell Enhancer Perturbation Assay.

Author information

1
Department of Obstetrics and Gynecology, Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, Dallas, TX, USA.
2
Department of Obstetrics and Gynecology, Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, Dallas, TX, USA. Gary.Hon@UTSouthwestern.edu.

Abstract

Transcriptional enhancers drive cell-type-specific gene expression patterns, and thus play key roles in development and disease. Large-scale consortia have extensively cataloged >one million putative enhancers encoded in the human genome. But few enhancers have been endogenously tested for function. For almost all enhancers, it remains unknown what genes they target and how much they contribute to target gene expression. We have previously developed a method called Mosaic-seq, which enables the high-throughput interrogation of enhancer activity by performing pooled CRISPRi-based epigenetic suppression of enhancers with a single-cell transcriptomic readout. Here, we describe an optimized version of this method, Mosaic-seq2. We have made several key improvements that have significantly simplified the library preparation process and increased the overall sensitivity and throughput of the method.

KEYWORDS:

CRISPRi; Enhancer; Single-cell RNA-seq; Single-cell perturbation

Supplemental Content

Full text links

Icon for Springer
Loading ...
Support Center