Genetic interaction mapping and exon-resolution functional genomics with a hybrid Cas9–Cas12a platform

  title={Genetic interaction mapping and exon-resolution functional genomics with a hybrid Cas9–Cas12a platform},
  author={Thomas Gonatopoulos-Pournatzis and Michael Aregger and Kevin R. Brown and Shaghayegh Farhangmehr and Ulrich Braunschweig and Henry N. Ward and Kevin C. H. Ha and Alexander Weiss and Maximilian Billmann and Tanja Durbic and Chad L. Myers and Benjamin J. Blencowe and Jason Moffat},
  journal={Nature Biotechnology},
Systematic mapping of genetic interactions (GIs) and interrogation of the functions of sizable genomic segments in mammalian cells represent important goals of biomedical research. To advance these goals, we present a CRISPR (clustered regularly interspaced short palindromic repeats)-based screening system for combinatorial genetic manipulation that employs coexpression of CRISPR-associated nucleases 9 and 12a (Cas9 and Cas12a) and machine-learning-optimized libraries of hybrid Cas9–Cas12a… 

Application of CHyMErA Cas9-Cas12a combinatorial genome-editing platform for genetic interaction mapping and gene fragment deletion screening.

CHyMErA (Cas hybrid for multiplexed editing and screening applications), which is based on the co-expression of Cas9 and Cas12a nucleases in conjunction with a hybrid guide RNA (hgRNA) engineered by the fusion of Cas 9 and Cas 12a guides and expressed from a single U6 promoter, is developed.

High-performance CRISPR-Cas12a genome editing for combinatorial genetic screening

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Multiplex enCas12a screens show functional buffering by paralogs is systematically absent from genome-wide CRISPR/Cas9 knockout screens

It is suggested that paralogs represent a targetable set of genetic dependencies that are systematically under-represented among cell-essential genes due to genetic buffering in monogenic CRISPR-based mammalian functional genomics approaches.

Multiplex enCas12a screens detect functional buffering among paralogs otherwise masked in monogenic Cas9 knockout screens

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Combinatorial CRISPR screening reveals functional buffering in autophagy

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Discovery of synthetic lethal and tumor suppressive paralog pairs in the human genome

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A High-Resolution Genome-Wide CRISPR/Cas9 Viability Screen Reveals Structural Features and Contextual Diversity of the Human Cell-Essential Proteome

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The optimized TKOv3 library, combined with the CEG2 reference set, provide an efficient, highly optimized platform for performing and assessing gene knockout screens in human cell lines and are demonstrated to be high effectiveness relative to reference sets of essential and nonessential genes.

Sequence determinants of improved CRISPR sgRNA design.

This work derived a new sequence model for predicting sgRNA efficiency in CRISPR/Cas9 knockout experiments and suggested new features including a preference for cytosine at the cleavage site that facilitate the genome-wide design of improved sg RNA for both knockout and CRISpri/a studies.

Genome-Scale CRISPR-Cas9 Knockout Screening in Human Cells

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Optimization of AsCas12a for combinatorial genetic screens in human cells

Improved Cas12a variants and sgRNA design rules enhance genome-wide screens and are validated by screening for synthetic lethalities in OVCAR8 and A375 cancer cells, discovering an interaction between MARCH5 and WSB2.