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ABSTRACT: Base-editing Screen

PROVIDER: PRJNA1265691 | ENA |

REPOSITORIES: ENA

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Dataset's files

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			Action	DRS
	SRR33677907_1.fastq.gz	Fastqsanger.gz
	SRR33677907_2.fastq.gz	Fastqsanger.gz
	SRR33677735_1.fastq.gz	Fastqsanger.gz
	SRR33677735_2.fastq.gz	Fastqsanger.gz
	SRR33677965_1.fastq.gz	Fastqsanger.gz

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Project description:Base editing screen in S.cerevisiae

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Whole-genome sequencing of Saccharomyces cerevisiae CEN.PK2-1C subjected to mtDNA base editing or Mito-BE screen

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Enhanced genome-wide knockout screens in bacteria with CRISPR base editors

Project description:Genome-wide knockout or knockdown screens have become powerful tools for the investigation of genotype-to-phenotype relationships. In bacteria, these screens commonly rely on transcriptional repression by dCas9, gene knockouts through Cas9 editing or random transposon mutagenesis, but depending on the technique, suffer from incomplete gene silencing, low editing efficiencies or they require massive library sizes. Here, we take a distinct approach with base editing to introduce premature stop codons or mutate start codons in Escherichia coli using a ScCas9 nickase derived base editor (ScBE3) that exhibits flexible PAM recognition. We then derive guide design rules by applying machine learning to a gene essentiality screen conducted in E. coli. For further improvement, we combined base-editing with Cas9-induced cleavage of the unedited cell fraction. The efficiency of this dual system was validated through a screen of conditionally essential E. coli genes. This improved setup that decouples the gene editing from the screening leads to more efficient guide depletion and confirmed previously published conditionally essential genes. Overall, base editing represents a useful tool for genome-wide knockout screens in bacteria and will eventually enable genome-wide knockout screens in a broader range of bacterial species to study their diverse genetics.

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Project description:Base editing improvement

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Enhanced base editing by co-expression of free uracil DNA glycosylase inhibitor

Project description:Even the latest generation of base editor (BE3) causes unwanted indels and non-C-to-T substitutions, compromising the fidelity of base editing outcome. Here we report a enhanced base editing system. The enhanced base edting system decreased the formation of unintended indels and C-to-A/C-to-G substitutions, and increased the frequency of desired C-to-T substitution, thereby improving both the accuracy and efficiency of base editing.

2017-08-29 | GSE98685 | GEO

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