Imperial College London
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ProfessorPaulFreemont

Faculty of Medicine, Department of Infectious Disease

Chair in Protein Crystallography
 
 
 
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Contact

 

+44 (0)20 7594 5327p.freemont

 
 
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Location

 

259Sir Alexander Fleming BuildingSouth Kensington Campus

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Summary

 

Publications

Citation

BibTex format

@article{Crone:2022,
author = {Crone, M and MacDonald, J and Freemont, P and Siciliano, V},
journal = {npj Systems Biology and Applications},
pages = {1--7},
title = {gDesigner: computational design of synthetic gRNAs for Cas12a-based transcriptional repression in mammalian cells},
url = {https://www.nature.com/articles/s41540-022-00241-w},
volume = {8},
year = {2022}
}
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RIS format (EndNote, RefMan)

TY  - JOUR
AB  - Synthetic networks require complex intertwined genetic regulation often relying on transcriptional activation or repression of target genes. CRISPRi-based transcription factors facilitate the programmable modulation of endogenous or synthetic promoter activity and can be aided by using software to select appropriate gRNAs and limit non-specific gene modulation. Here, we develop a computational software pipeline, gDesigner, that enables the automated selection of orthogonal gRNAs with minimized off-target effects and promoter crosstalk. We next engineered a  Lachnospiraceae bacterium Cas12a (dLbCas12a)-based repression system that downregulates target gene expression by means of steric hindrance of the cognate promoter. Finally, we generated a library of orthogonal synthetic dCas12a-repressed promoters and experimentally demonstrated it in HEK293FT, U2OS and H1299 cells lines. Our system expands the toolkit of mammalian synthetic promoters with a new complementary and orthogonal CRISPRi-based system, ultimately enabling the design of synthetic promoter libraries for multiplex gene perturbation that facilitate the understanding of complex cellular phenotypes.
AU  - Crone,M
AU  - MacDonald,J
AU  - Freemont,P
AU  - Siciliano,V
EP  - 7
PY  - 2022///
SN  - 2056-7189
SP  - 1
TI  - gDesigner: computational design of synthetic gRNAs for Cas12a-based transcriptional repression in mammalian cells
T2  - npj Systems Biology and Applications
UR  - https://www.nature.com/articles/s41540-022-00241-w
UR  - http://hdl.handle.net/10044/1/99498
VL  - 8
ER  -
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