A high-content flow cytometry and dual CRISPR-Cas9 based platform to quantify genetic interactions

Natasha Ramakrishnan, Taylor Malachowski, Priyanka Verma

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

1 Scopus citations


Probing epistasis between two genes can be a critical first step in identifying the molecular players in a cellular pathway. The advent of CRISPR-Cas mediated genetic screen has enabled studying of these genetic interactions at a genomic scale. However, when combining depletion of two genes using CRISPR Cas9, reduced targeting efficiencies due to competition for Cas loading and recombination in the cloning step have emerged as key challenges. Moreover, given conventional CRISPR screens typically involve comparison between the initial and final time point, it is difficult to parse the time kinetics with which a perturbed genetic interaction impacts viability, and it also becomes challenging to assess epistasis with essential genes. Here, we discuss a high-throughput flow-based approach to study genetic interactions. By utilizing two different Cas9 orthologs and monitoring viability at multiple time points, this approach helps to effectively mitigate the limitations of Cas9 competition and enables assessment of genetic interactions with both essential and non-essential genes at a high temporal resolution.

Original languageEnglish
Title of host publicationMethods in Cell Biology
PublisherAcademic Press Inc.
Number of pages14
ISBN (Print)9780443188985
StatePublished - Jan 2024

Publication series

NameMethods in Cell Biology
ISSN (Print)0091-679X


  • DNA repair
  • GFP competition
  • High-content flow cytometry
  • Replication
  • SaCas9


Dive into the research topics of 'A high-content flow cytometry and dual CRISPR-Cas9 based platform to quantify genetic interactions'. Together they form a unique fingerprint.

Cite this