A dual sgRNA library design to probe genetic modifiers using genome-wide CRISPRi screens

Alina Guna; Katharine R Page; Joseph M Replogle; Theodore K Esantsi; Maxine L Wang; Jonathan S Weissman; Rebecca M Voorhees

doi:10.1101/2023.01.22.525086

A dual sgRNA library design to probe genetic modifiers using genome-wide CRISPRi screens

bioRxiv [Preprint]. 2023 Aug 14:2023.01.22.525086. doi: 10.1101/2023.01.22.525086.

Authors

Alina Guna^{1

2}, Katharine R Page¹, Joseph M Replogle^{2

3

4}, Theodore K Esantsi^{2

4}, Maxine L Wang^{1

2}, Jonathan S Weissman^{2

4

5

6}, Rebecca M Voorhees¹

Affiliations

¹ Division of Biology and Biological Engineering, California Institute of Technology, 1200 E. California Ave., Pasadena, CA 91125, USA.
² Whitehead Institute for Biomedical Research, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
³ Medical Scientist Training Program, University of California, San Francisco, San Francisco, CA 94158, USA.
⁴ Howard Hughes Medical Institute, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
⁵ Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
⁶ David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA,02142, USA.

Abstract

Mapping genetic interactions is essential for determining gene function and defining novel biological pathways. We report a simple to use CRISPR interference (CRISPRi) based platform, compatible with Fluorescence Activated Cell Sorting (FACS)-based reporter screens, to query epistatic relationships at scale. This is enabled by a flexible dual-sgRNA library design that allows for the simultaneous delivery and selection of a fixed sgRNA and a second randomized guide, comprised of a genome-wide library, with a single transduction. We use this approach to identify epistatic relationships for a defined biological pathway, showing both increased sensitivity and specificity than traditional growth screening approaches.

Publication types

Preprint

Abstract

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