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Related Concept Videos

Genetic Screens02:46

Genetic Screens

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Genetic screens are tools used to identify genes and mutations responsible for phenotypes of interest. Genetic screens help identify individuals or a group of people at risk of developing  genetic diseases and help them with early intervention, targeted therapy, and reproductive options.
Forward genetic screens
Forward or “classical” genetic screens involve creating random mutations in an organism’s DNA using radiation, mutagens, or insertion of additional bases, which...
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Protocol for Comprehensive Synthetic Lethality Screens.

Damia Romero-Moya1, Jeroen P Roose1,2,3

  • 1Department of Anatomy, University of California San Francisco (UCSF), 513 Parnassus Avenue, Room HSW-1326, San Francisco, CA 94143-0452, USA.

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|September 28, 2020
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Summary

This study details a protocol for synthetic lethality screening in leukemia cells, using a PI3K inhibitor and gene depletion to identify cell death triggers. The method is adaptable for researchers investigating cancer therapeutics.

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Area of Science:

  • Cell Biology
  • Molecular Biology
  • Cancer Research

Background:

  • Synthetic lethality is a promising strategy for targeted cancer therapy.
  • Identifying synthetic lethal interactions requires robust screening methods.
  • Jurkat T cell leukemia provides a model for studying T cell malignancies.

Purpose of the Study:

  • To provide a detailed protocol for synthetic lethality screening in Jurkat T cells.
  • To establish a method for measuring the combinatorial effects of PI3K inhibition and gene depletion.
  • To facilitate the adaptation of this protocol by other investigators.

Main Methods:

  • Utilized an ultra-complex shRNA library for comprehensive gene depletion.
  • Employed cell death as the primary readout for synthetic lethality.
  • Combined a pan-PI3K inhibitor (GDC0941) with specific gene depletion.
  • Detailed protocol steps, coverage considerations, time frames, and potential bottlenecks.

Main Results:

  • Successfully established a protocol for synthetic lethality screening in a leukemia cell line.
  • Demonstrated the measurement of combinatorial drug and gene-knockdown effects.
  • Provided practical insights and troubleshooting for protocol execution.

Conclusions:

  • The described protocol is readily adaptable for researchers in cancer and molecular biology.
  • This methodology aids in discovering novel synthetic lethal interactions for therapeutic development.
  • The protocol facilitates efficient screening of drug-gene interactions in leukemia models.