An optimized genome-wide, virus-free CRISPR screen for mammalian cells.
Journal
Cell reports methods
ISSN: 2667-2375
Titre abrégé: Cell Rep Methods
Pays: United States
ID NLM: 9918227360606676
Informations de publication
Date de publication:
23 08 2021
23 08 2021
Historique:
entrez:
22
12
2021
pubmed:
23
12
2021
medline:
23
12
2021
Statut:
ppublish
Résumé
Pooled CRISPR screens have been widely applied to mammalian and other organisms to elucidate the interplay between genes and phenotypes of interest. The most popular method for delivering the CRISPR components into mammalian cells is lentivirus based. However, because lentivirus is not always an option, virus-free protocols are starting to emerge. Here, we demonstrate an improved virus-free, genome-wide CRISPR screening platform for Chinese hamster ovary cells with 75,488 gRNAs targeting 15,028 genes. Each gRNA expression cassette in the library is precisely integrated into a genomic landing pad, resulting in a very high percentage of single gRNA insertions and minimal clonal variation. Using this platform, we perform a negative selection screen on cell proliferation that identifies 1,980 genes that affect proliferation and a positive selection screen on the toxic endoplasmic reticulum stress inducer, tunicamycin, that identifies 77 gene knockouts that improve survivability.
Identifiants
pubmed: 34935002
doi: 10.1016/j.crmeth.2021.100062
pmc: PMC8687118
mid: NIHMS1763636
pii:
doi:
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Research Support, N.I.H., Extramural
Langues
eng
Subventions
Organisme : NIGMS NIH HHS
ID : R35 GM119850
Pays : United States
Commentaires et corrections
Type : ErratumIn
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