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Related Experiment Video

Updated: Nov 20, 2025

Pooled CRISPR-Based Genetic Screens in Mammalian Cells
09:05

Pooled CRISPR-Based Genetic Screens in Mammalian Cells

Published on: September 4, 2019

22.7K

High-content imaging-based pooled CRISPR screens in mammalian cells.

Xiaowei Yan1, Nico Stuurman1, Susana A Ribeiro1,2

  • 1Department of Cellular and Molecular Pharmacology, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA.

The Journal of Cell Biology
|January 19, 2021
PubMed
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This study introduces optical enrichment, a microscopy-based method for pooled CRISPR screens. It enables automated identification and isolation of cells with specific CRISPR-induced phenotypes, advancing genetic screening capabilities.

Area of Science:

  • Molecular Biology
  • Genetics
  • Cell Biology

Background:

  • CRISPR gene inactivation links genes to cellular phenotypes.
  • Pooled CRISPR screens using sgRNAs are limited to selectable phenotypes.
  • A need exists for screening phenotypes not amenable to traditional selection methods.

Purpose of the Study:

  • To develop a microscopy-based approach for pooled CRISPR screens.
  • To enable the selection of cells based on visually identifiable CRISPR-induced phenotypes.
  • To expand the scope of pooled CRISPR screening beyond chemical or FACS enrichment.

Main Methods:

  • Developed 'optical enrichment,' a microscopy-based screening method.
  • Automated image analysis identifies cells with specific CRISPR phenotypes.

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Last Updated: Nov 20, 2025

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  • Photoactivation marks selected cells for isolation via fluorescence-activated cell sorting (FACS).
  • A μManager plugin facilitates automated cell identification and photoactivation.

    • Main Results:

    • Screened ~1.5 million cells in 8 hours.
    • Successfully screened 6,092 sgRNAs targeting 544 genes for nuclear size regulation.
    • Identified 14 bona fide genes affecting nuclear size.

    Conclusions:

    • Optical enrichment provides a scalable method for imaging-based pooled CRISPR screens.
    • This approach significantly expands the types of phenotypes that can be screened using CRISPR.
    • Facilitates high-throughput genetic screens for visually defined cellular characteristics.