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Targeted modulation of chicken genes in vitro using CRISPRa and CRISPRi toolkit

ABSTRACT: Targeted modulation of chicken genes in vitro using CRISPRa and CRISPRi toolkit

PROVIDER: PRJNA898374 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR22195292_1.fastq.gz	Fastqsanger.gz
	SRR22195292_2.fastq.gz	Fastqsanger.gz
	SRR22195293_1.fastq.gz	Fastqsanger.gz
	SRR22195293_2.fastq.gz	Fastqsanger.gz
	SRR22195294_1.fastq.gz	Fastqsanger.gz

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Targeted Modulation of Chicken Genes In Vitro Using CRISPRa and CRISPRi Toolkit.

Project description:Engineering of clustered regularly interspaced short palindromic repeats (CRISPR) and the CRISPR-associated protein 9 (Cas9) system has enabled versatile applications of CRISPR beyond targeted DNA cleavage. Combination of nuclease-deactivated Cas9 (dCas9) and transcriptional effector domains allows activation (CRISPRa) or repression (CRISPRi) of target loci. To demonstrate the effectiveness of the CRISPR-mediated transcriptional regulation in chickens, three CRISPRa (VP64, VPR, and p300) and three CRISPRi (dCas9, dCas9-KRAB, and dCas9-KRAB-MeCP2) systems were tested in chicken DF-1 cells. By introducing guide RNAs (gRNAs) targeting near the transcription start site (TSS) of each gene in CRISPRa and CRISPRi effector domain-expressing chicken DF-1 cell lines, significant gene upregulation was induced in dCas9-VPR and dCas9-VP64 cells, while significant downregulation was observed with dCas9 and dCas9-KRAB. We further investigated the effect of gRNA positions across TSS and discovered that the location of gRNA is an important factor for targeted gene regulation. RNA sequencing analysis of IRF7 CRISPRa and CRISPRi- DF-1 cells revealed the specificity of CRISPRa and CRISPRi-based targeted transcriptional regulation with minimal off-target effects. These findings suggest that the CRISPRa and CRISPRi toolkits are an effective and adaptable platform for studying the chicken genome by targeted transcriptional modulation.

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Targeted modulation of chicken genes in vitro using CRISPRa and CRISPRi toolkit

Project description:To investigate the effectiveness of CRISPR based gene modulation, IRF7 was repressed in dCas9-KRAB DF1 cell line or activated in dCas9-VPR DF1 cell line followed by whole transcriptome analysis

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