Keratinocyte CRISPR screens
Ontology highlight
ABSTRACT:
PROVIDER: EGAS00001002714 | EGA |
REPOSITORIES: EGA
Similar Datasets
Project description:Immortalised HaCaT keratinocytes were transduced with Cas9 and the CRISPR-KO v1.1 genome-wide gRNA library. The gRNA library was prepared from genomic DNA isolated 14 days post library transduction. gRNA representation will be compared to the original CRISPR-KO v1.1 library to reveal genes essential for HaCaT survival and growth. .
This dataset contains all the data available for this study on 2019-08-14.
| EGAD00001005252 | EGA
Project description:Affymetrix SNP arrays were performed according to the manufacturer's directions on DNA extracted from human skin keratinocytes derived from healthy donors, as well as from telomerase immortalised human skin keratinocytes (N/TERT), transduced with mock, EGFP, or EGFP-GLI2DeltaN transgenes to compare chromosomal ploidy status.
2012-02-28 | GSE36105 | GEO
Project description:Affymetrix SNP arrays were performed according to the manufacturer's directions on DNA extracted from human skin keratinocytes derived from healthy donors, as well as from telomerase immortalised human skin keratinocytes (N/TERT), transduced with mock, EGFP, or EGFP-GLI2DeltaN transgenes to compare chromosomal ploidy status. Affymetrix SNP arrays were performed according to the manufacturer's directions on DNA extracted from human skin keratinocytes derived from healthy donors, as well as from telomerase immortalised human skin keratinocytes (N/TERT) to compare chromosomal ploidy status
2012-02-28 | E-GEOD-36105 | biostudies-arrayexpress
Project description:Genome-wide CRISPR screens were performed in Huh-7 hepatoma cell line to identify genes regulating ferroptosis sensitivity. Huh-7 cells were transduced with Human GeCKO library and selected by RSL-3.
2023-04-17 | GSE228883 | GEO
Project description:To identify genes for radioresistance in lung cancer, unbiased CRISPR/Cas9 library knockout screening was performed in lung cancer containing a pooled 903 post-translational modification (PTM) related regulatory proteins library. Two populations of cells were either nonirradiated or irradiated at 9 Gy, and they were cultured in vitro. Then genomic DNA was acquired to readout the sgRNA representation by deep sequencing.
2026-02-25 | GSE303409 | GEO
Project description:This study aimed to enhance the therapeutic effect of MSC-derived extracellular vesicles (EVs) by overexpressing the TSG-6 gene using CRISPR activation and evaluate the biological activity of these modified EVs on human intervertebral disc (IVD) cells in vitro. n immortalized human MSC line was transduced with a CRISPR activation system targeting TSG-6. MSC-EVs were collected via ultracentrifugation, and their particle size/number was analyzed. Gene and protein expression were assessed to evaluate transduction efficiency. EV proteomics was analyzed by mass spectrometry. Human IVD cells were isolated from patients undergoing spinal surgery, pre-stimulated with IL-1β, and treated with MSC-EVs.
2025-05-07 | PXD060381 | Pride
Project description:Here, we introduce CRaft-ID (CRISPR-based microRaft followed by gRNA identification) to enable the screening of image-based phenotypes such as subcellular protein localization from a pool of lenti-CRISPR bulk infected cells. Clonal colonies selected from microRaft arrays are isolated after phenotyping for a targeted-sequencing library preparation to identify the infected sgRNA in each colony. sgRNA representation in the pool of cells is measured with a targeted-sequencing library preparation from the bulk sample.
2019-11-02 | GSE139815 | GEO
Project description:CRISPR–Cas9 screening relies on uniform representation of single-guide RNA (sgRNA) libraries to enable accurate gene discovery. However, technical biases during library preparation can compromise screen performance. Here we show that commonly used “all-in-one” CRISPR vectors expressing both Cas9 and sgRNAs drive unintended Cas9 protein expression in Escherichia coli during plasmid amplification. This bacterial Cas9 expression causes guide-specific toxicity, leading to selective loss of sgRNAs and highly skewed library representation. We demonstrate that this effect occurs across multiple bacterial strains and affects both targeted and genome-wide libraries, including widely used human CRISPR libraries. Mechanistically, toxicity is driven by Cas9 expression rather than plasmid size and is only partially alleviated by catalytically inactive Cas9. Importantly, replacing the EF-1α promoter with a mouse phosphoglycerate kinase promoter suppresses Cas9 expression in bacteria while preserving genome editing efficiency in mammalian cells and restoring sgRNA uniformity. These findings identify a previously unrecognized source of bias in CRISPR library preparation and provide a practical solution to improve screening fidelity.
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Project description:H1-HeLa cells were stably transduced with lentiCas9-Blast (Addgene, Plasmid #52962) and subsequently selected using blasticidin to generate constitutively expressing Cas9 H1-HeLa cells. A single Cas9-expressing H1-HeLa clone was then transduced with lentivirus without a selection marker to stably express CDHR3 C529Y (H1-HeLa+CDHR3). A single CDHR3-expressing H1-HeLa clone was then chosen based on RT-qPCR of CHDR3 expression and RV-C15 RNA levels for mutagenesis. 300 million of the H1-HeLa cells constitutively expressing CDHR3 and Cas9 were transduced with the lentiGuide-Puro from the GeCKO v2 library at a MOI of 0.3. Cells were selected using puromycin and heterogeneous H1-HeLa knockout cell populations were subsequently pooled together. The CRISPR genetic screens were started 10 days post transduction. >1000-fold coverage of mutagenized cells (libraries A and B) was infected with either RV-C15 (MOI=1 PFU/cell) or EV-D68 Missouri (MOI=1 PFU/cell). RV-C15 infection was repeated for an additional round at 6 days post-infection. As soon as appearance of visibly viable colonies was observed, populations of virus-resistant cells were pooled and harvested. Uninfected starting populations of mutagenized cells were used as the unselected reference. Total genomic DNA from both virus-resistant and uninfected cells was respectively extracted using QIAamp DNA Mini Kit (Qiagen). The inserted guide RNA sequences were retrieved from the genomic DNA by PCR amplification. The PCR products were then purified and subjected to NextSeq platform (Illumina) next-generation sequencing.
2019-07-10 | E-MTAB-8125 | biostudies-arrayexpress
Project description:CRISPR–Cas9 screening relies on uniform representation of single-guide RNA (sgRNA) libraries to enable accurate gene discovery. However, technical biases during library preparation can compromise screen performance. Here we show that commonly used “all-in-one” CRISPR vectors expressing both Cas9 and sgRNAs drive unintended Cas9 protein expression in Escherichia coli during plasmid amplification. This bacterial Cas9 expression causes guide-specific toxicity, leading to selective loss of sgRNAs and highly skewed library representation. We demonstrate that this effect occurs across multiple bacterial strains and affects both targeted and genome-wide libraries, including widely used human CRISPR libraries. Mechanistically, toxicity is driven by Cas9 expression rather than plasmid size and is only partially alleviated by catalytically inactive Cas9. Importantly, replacing the EF-1α promoter with a mouse phosphoglycerate kinase promoter suppresses Cas9 expression in bacteria while preserving genome editing efficiency in mammalian cells and restoring sgRNA uniformity. These findings identify a previously unrecognized source of bias in CRISPR library preparation and provide a practical solution to improve screening fidelity.
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