CRISPR screens are feasible in TP53 wild-type cells
Ontology highlight
ABSTRACT: CRISPR-Cas9 genome-wide screens were performed in retinal pigment epithelial cells (RPE1) with either wild-type TP53 gene, or a TP53-null background. Results show wild-type TP53 has minimal impact on the efficiency of CRISPR dropout screens.
Project description:This is data for the evaluation of a new way of counting sgRNAs in CRISPR screens using padlock probes and UMIs. It is compared to the typical PCR-based approach. In particular, a dropout screen was performed in MiaPaCa-2 cells using the Human Kinome CRISPR pooled library (Addgene #75314)
Project description:To compare the impact of several TP53 mutant variants in an isogenic setting, different TP53 mutations were introduced in HCT116 colorectal carcinoma cells. This parental cell line is wild-type for TP53 and shows a prototypical p53 response. To ensure unambigous genotype-phenotype correlations, the cell were haploidized prior to CRISPR-editing by introducing inactivating deletions of intronic splicing into one of the two TP53 alleles, leaving only one functional copy of TP53. The remaining TP53 allele was altered by inserting a LoxP-flanked transcriptional stop cassette (Lox-Stop-Lox, LSL) into intron 4, which allowed reversible silencing of TP53 expression. The LSL cassette was then specifically targeted with CRISPR/Cas9 to introduce a variety of different mutant p53 alleles. The competency of the mutated p53 allele to induce a p53 response upon activation using Nutlin-3a was then assessed in an RNAseq experiment.
Project description:We performed CRISPR screens on both a sub-library and a genome-wide scale in human intestinal organoids to discover cancer driver genes. We investigated the Wnt and the TGFB pathway and used both WT, APC-mutant and APC-TP53-mutant organoids.
Project description:Comparison of the TP53 wild-type myeloma cell line AMO1 with the CRISPR/Cas9 engineered AMO1 cell line named UMC901. UMC901 harbors bi-allelic alterations to TP53: TP53 del/mut. Analysis of impact of TP53 alterations on gene transcription and identification of affected pathways by transcriptome-wide differential gene expression analysis.
Project description:In order to investigate the potential role of TP53 in regulating translation, HCT116 wild type and TP53 knockout cells were analyzed using both RNA sequencing, Ribosome sequencing and nascent proteome analysis. The cells were treated with 0.2 µg/ml Neocarcinostatin (NCS) to induce DNA damage and activate TP53.
Project description:We performed CRISPR screens on both a sub-library and a genome-wide scale in human intestinal organoids to discover cancer driver genes. We investigated the Wnt and the TGFB pathway and used both WT, APC-mutant and APC-TP53-mutant organoids.
Project description:We performed CRISPR screens on both a sub-library and a genome-wide scale in human intestinal organoids to discover cancer driver genes. We investigated the Wnt and the TGFB pathway and used both WT, APC-mutant and APC-TP53-mutant organoids.
Project description:We performed CRISPR screens on both a sub-library and a genome-wide scale in human intestinal organoids to discover cancer driver genes. We investigated the Wnt and the TGFB pathway and used both WT, APC-mutant and APC-TP53-mutant organoids.