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Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system期刊论文

作者: Albert W Cheng Haoyi Wang Hui Yang Linyu Shi Yarden Katz Thorold W Theunissen Sudharshan Rangarajan Chikdu S Shivalila Daniel B Dadon Rudolf Jaenisch
DOI:10.1038/cr.2013.122

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页码: 1163-1171
被引频次: 314
出版者: INST BIOCHEMISTRY & CELL BIOLOGY,Nature Publishing Group
ISSN: 1001-0602
卷期: Volume:23    Issue:10
语言: English
摘要: Technologies allowing for specific regulation of endogenous genes are valuable for the study of gene functions and have great potential in therapeutics. We created the CRISPR-on system, a two-component transcriptional activator consisting of a nuclease-dead Cas9 (dCas9) protein fused with a transcriptional activation domain and single guide RNAs (sgRNAs) with complementary sequence to gene promoters. We demonstrate that CRISPR-on can efficiently activate exogenous reporter genes in both human and mouse cells in a tunable manner. In addition, we show that robust reporter gene activation in vivo can be achieved by injecting the system components into mouse zygotes. Furthermore, we show that CRISPR-on can activate the endogenous IL1RN, SOX2, and OCT4 genes. The most efficient gene activation was achieved by clusters of 3-4 sgRNAs binding to the proximal promoters, suggesting their synergistic action in gene induction. Significantly, when sgRNAs targeting multiple genes were simultaneously introduced into cells, robust multiplexed endogenous gene activation was achieved. Genome-wide expression profiling demonstrated high specificity of the system.
相关主题: 转录激活, 表达谱分析, RNA, 报告基因, 内源基因, 激活系统, 基因功能, 引导, CRISPR, synthetic biology, transcription factor, artificial transcription factor, gene expression, BACTERIA, ARCHAEA, TAL EFFECTORS, CAS SYSTEMS, EXPRESSION, BINDING, CELL BIOLOGY, NIH 3T3 Cells, Promoter Regions, Genetic, RNA, Guide - genetics, Deoxyribonuclease I - genetics, Humans, Mice, Inbred C57BL, Transcriptional Activation, Transcription Factors - genetics, Animals, Cloning, Molecular, Clustered Regularly Interspaced Short Palindromic Repeats, HEK293 Cells, Recombinant Fusion Proteins - genetics, Female, Mice, HeLa Cells, Transgenes, Index Medicus, Original,

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