Direct-seq: programmed gRNA scaffold for streamlined scRNA-seq in CRISPR screen.

Genome Biology
Qingkai SongLijia Ma

Abstract

CRISPR-based genome perturbation provides a new avenue to conveniently change DNA sequences, transcription, and epigenetic modifications in genetic screens. However, it remains challenging to assay the complex molecular readouts after perturbation at high resolution and at scale. By introducing an A/G mixed capture sequence into the gRNA scaffold, we demonstrate that gRNA transcripts could be directly reverse transcribed by poly (dT) primer together with the endogenous mRNA, followed by high-content molecular phenotyping in scRNA-seq (Direct-seq). With this method, the CRISPR perturbation and its transcriptional readouts can be profiled together in a streamlined workflow.

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Citations

Feb 25, 2021·ACS Synthetic Biology·Xin Yi ChooOwen J L Rackham

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Datasets Mentioned

BETA
GSE146194

Methods Mentioned

BETA
RNA-seq
scRNA-seq
PCR
CROP-seq
gene
fluorescence-activated cell sorting
flow cytometry
single-cell sequencing
Chip
HTseq

Software Mentioned

Fluidigm C1
Seurat
TIDE
Cell Ranger Single - Cell
R package clusterProfiler
HTseq

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