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Optimized CRISPR-Cas12a platform for combinatorial drop-out genetic screening (RNA-seq)

ABSTRACT: Optimized CRISPR-Cas12a platform for combinatorial drop-out genetic screening (RNA-seq)

PROVIDER: PRJNA592183 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR10546800.fastq.gz	Fastqsanger.gz
	SRR10546801.fastq.gz	Fastqsanger.gz
	SRR10546802.fastq.gz	Fastqsanger.gz
	SRR10546803.fastq.gz	Fastqsanger.gz
	SRR10546804.fastq.gz	Fastqsanger.gz

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Optimized CRISPR-Cas12a platform for combinatorial drop-out genetic screening (RNA-seq)

Project description:Combinatorial genetic perturbations have been utilized to identify synthetic sick/lethal genetic interactions for cancer drug target discovery. Current methods for high-throughput combinatorial genetic screening are inefficient and cumbersome. Here we developed a simple, robust, and high-performance CRISPR-Cas12a-based approach for unbiased, combinatorial genetic screening in cancer cells.

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Project description:Systematic mapping of genetic interactions and interrogation of the functions of sizeable genomic segments in mammalian cells represent important goals of biomedical research. To advance these goals, we present a CRISPR-based screening system for combinatorial genetic manipulation that employs co-expression of Cas9 and Cas12a nucleases and machine learning-optimized libraries of hybrid Cas9-Cas12a guide RNAs. This system, named CHyMErA (Cas Hybrid for Multiplexed Editing and Screening Applications), outperforms genetic screens using Cas9 or Cas12a editing alone. Application of CHyMErA to the ablation of mammalian paralog gene pairs reveals extensive genetic interactions and uncovers phenotypes normally masked by functional redundancy. Application of CHyMErA in a chemo-genetic interaction screen identifies genes that impact cell growth in response to mTOR pathway inhibition. Moreover, by systematically targeting thousands of alternative splicing events, CHyMErA identifies exons underlying human cell line fitness. CHyMErA thus represents an effective screening approach for genetic interaction mapping and the functional analysis of sizeable genomic regions, such as alternative exons.

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