A microRNA-inducible CRISPR-Cas9 platform serves as a microRNA sensor and cell-type-specific genome regulation tool

Nature Cell Biology
Xi-Wen WangYangming Wang

Abstract

microRNAs (miRNAs) are small noncoding RNAs that play important regulatory roles in plants, animals and viruses. Measuring miRNA activity in vivo remains a big challenge. Here, using an miRNA-mediated single guide RNA (sgRNA)-releasing strategy and dCas9-VPR to drive a transgene red fluorescent protein, we create an miRNA sensor that can faithfully measure miRNA activity at cellular levels and use it to monitor differentiation status of stem cells. Furthermore, by designing sgRNAs to target endogenous loci, we adapted this system to control the expression of endogenous genes or mutate specific DNA bases upon induction by cell-type-specific miRNAs. Finally, by miRNA sensor library screening, we discover a previously undefined layer of heterogeneity associated with miR-21a activity in mouse embryonic stem cells. Together, these results highlight the utility of an miRNA-induced CRISPR-Cas9 system as miRNA sensors and cell-type-specific genome regulation tools.

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Jan 11, 2020·The Plant Journal : for Cell and Molecular Biology·Paul D FraserAlisdair R Fernie
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Methods Mentioned

BETA
flow cytometry
transfection
PCR
immunoprecipitation
amplicon sequencing
electrophoresis

Software Mentioned

BE
DESeq
FLASH
ON
TargetScan7
GSEA
MICR
Trimmomatic
R package DescTools
DAVID

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