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| Status |
Public on Feb 28, 2019 |
| Title |
Transcriptome profiling of H1-iCas9 cells grown on laminin and on MEFs |
| Organisms |
Homo sapiens; Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
This experiment is part of a study of differentially essential genes in H1 hPSCs grown on different substrates ("Essential gene profiles for human pluripotent stem cells identify uncharacterized genes and substrate dependencies", Mair et al., Cell Reports 2018). The cells used stably express DOX-inducible Cas9 in the AAVS1 locus and were profiled together with unmodified H1 wt hPCS on both substrates to asses transcriptional changes.
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| Overall design |
Human pluripotent stem cells (hPSCs) provide an invaluable tool for modeling diseases and hold therapeutic promise for regenerative medicine. Hence, there is great interest in understanding the genetic wiring that governs pluripotency and lineage differentiation. A critical first step involves a systematic understanding and cataloging of genes that are indispensable for hPSC maintenance and proliferation. In order to map core and context-specific genetic determinants of hPSC fitness, we generated an inducible Cas9 H1 hPSC line and performed genome-scale loss-of-function screens using a highly optimized CRISPR-Cas9 library to identify essential genes in hPSCs cultured on feeder cells and in feeder-free conditions. Dependencies that were predictable in hPSCs were identified such as FOXH1 and VENTX, genes that encode transcription factors that have been implicated in stem cell biology and differentiation, as well as genes not previously associated with stem cells, including an uncharacterized gene encoding a potential hPSC regulator, C22orf43/DRICH1. Genes found to be essential in hPSCs are substantially different from essential gene sets derived from a broad set of cell lines, and gene essentiality in hPSCs is highly context-dependent, i.e. essential gene sets vary with different substrates. We present condition-specific CRISPR screens in hPSCs, and establish parameters for genome-wide screens in this cell type. Ultimately, these data will facilitate the investigation of essential genes in hPSCs in different contexts and the characterization of unappreciated regulators of hPSC biology to understand the genetic wiring of hPSC fitness and pluripotency.
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| Contributor(s) |
Mair B, Brown KR, Moffat J |
| Citation(s) |
30970261 |
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| Submission date |
Jan 23, 2019 |
| Last update date |
Aug 14, 2020 |
| Contact name |
Jason Moffat |
| E-mail(s) |
j.moffat@utoronto.ca, jason.moffat@sickkids.ca
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| Phone |
4379965938
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| Organization name |
University of Toronto
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| Department |
Molecular Genetics
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| Street address |
686 Bay Street, Room 13.9705
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| City |
Toronto |
| State/province |
ON |
| ZIP/Postal code |
M5G0A4 |
| Country |
Canada |
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| Platforms (2) |
| GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
| GPL19057 |
Illumina NextSeq 500 (Mus musculus) |
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| Samples (17)
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| This SubSeries is part of SuperSeries: |
| GSE156298 |
Essential gene profiles for human pluripotent stem cells identify uncharacterized genes and substrate dependencies |
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| Relations |
| BioProject |
PRJNA516653 |
| SRA |
SRP181645 |
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