In this section
@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}
}
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
VL - 8
ER -