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Methods Mol Biol. 2016;1358:43-57. doi: 10.1007/978-1-4939-3067-8_3.

Transcriptional Regulation with CRISPR/Cas9 Effectors in Mammalian Cells.

Author information

1
Program in Molecular Medicine, Diabetes Center of Excellence, University of Massachusetts Medical School, 368 Plantation Street AS7-2057, Worcester, MA, 01605, USA.
2
Program in Molecular Medicine, Diabetes Center of Excellence, University of Massachusetts Medical School, 368 Plantation Street AS7-2057, Worcester, MA, 01605, USA. rene.maehr@umassmed.edu.

Abstract

CRISPR/Cas9-based regulation of gene expression provides the scientific community with a new high-throughput tool to dissect the role of genes in molecular processes and cellular functions. Single-guide RNAs allow for recruitment of a nuclease-dead Cas9 protein and transcriptional Cas9-effector fusion proteins to specific genomic loci, thereby modulating gene expression. We describe the application of a CRISPR-Cas9 effector system from Streptococcus pyogenes for transcriptional regulation in mammalian cells resulting in activation or repression of transcription. We present methods for appropriate target site selection, sgRNA design, and delivery of dCas9 and dCas9-effector system components into cells through lentiviral transgenesis to modulate transcription.

KEYWORDS:

CRISPR/Cas9; Cas9-effectors; Gene activation; Gene repression; Transcriptional regulation

PMID:
26463376
DOI:
10.1007/978-1-4939-3067-8_3
[Indexed for MEDLINE]

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