dCas9 regulator to neutralize competition in CRISPRi circuits.
Journal
Nature communications
ISSN: 2041-1723
Titre abrégé: Nat Commun
Pays: England
ID NLM: 101528555
Informations de publication
Date de publication:
16 03 2021
16 03 2021
Historique:
received:
03
11
2020
accepted:
29
01
2021
entrez:
17
3
2021
pubmed:
18
3
2021
medline:
24
3
2021
Statut:
epublish
Résumé
CRISPRi-mediated gene regulation allows simultaneous control of many genes. However, highly specific sgRNA-promoter binding is, alone, insufficient to achieve independent transcriptional regulation of multiple targets. Indeed, due to competition for dCas9, the repression ability of one sgRNA changes significantly when another sgRNA becomes expressed. To solve this problem and decouple sgRNA-mediated regulatory paths, we create a dCas9 concentration regulator that implements negative feedback on dCas9 level. This allows any sgRNA to maintain an approximately constant dose-response curve, independent of other sgRNAs. We demonstrate the regulator performance on both single-stage and layered CRISPRi-based genetic circuits, zeroing competition effects of up to 15-fold changes in circuit I/O response encountered without the dCas9 regulator. The dCas9 regulator decouples sgRNA-mediated regulatory paths, enabling concurrent and independent regulation of multiple genes. This allows predictable composition of CRISPRi-based genetic modules, which is essential in the design of larger scale synthetic genetic circuits.
Identifiants
pubmed: 33727557
doi: 10.1038/s41467-021-21772-6
pii: 10.1038/s41467-021-21772-6
pmc: PMC7966764
doi:
Substances chimiques
RNA, Guide
0
CRISPR-Associated Protein 9
EC 3.1.-
Types de publication
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Langues
eng
Sous-ensembles de citation
IM
Pagination
1692Références
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