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Related Concept Videos

CRISPR/Cas9 Genome Editing01:28

CRISPR/Cas9 Genome Editing

The CRISPR-Cas system serves as a bacterial defense mechanism against invading genetic elements such as viruses and plasmids, forming the foundation for its adaptation as a powerful genome-editing tool. Originally discovered in prokaryotes, this system has been repurposed to revolutionize genetic engineering across a wide range of organisms, including plants, animals, and humans. The core component, Cas9, is an endonuclease derived from Streptococcus pyogenes, capable of introducing...
Genetic Screens02:46

Genetic Screens

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 result in visible changes...
CRISPR01:59

CRISPR

Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced Short...
CRISPR01:59

CRISPR

Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced Short...
CRISPR and crRNAs02:53

CRISPR and crRNAs

Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...

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Related Experiment Video

Updated: Jun 16, 2026

Cell Surface Receptor Identification Using Genome-Scale CRISPR/Cas9 Genetic Screens
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CROPseq-multi: a universal solution for multiplexed perturbation in high-content pooled CRISPR screens.

Russell T Walton1,2, Yue Qin1,3, Paul C Blainey1

  • 1Broad Institute of MIT and Harvard, Cambridge, MA, USA.

Biorxiv : the Preprint Server for Biology
|April 1, 2024
PubMed
Summary

Researchers developed CROPseq-multi, a new lentiviral system for multiplexed genetic screening using Streptococcus pyogenes (Sp) Cas9. This tool efficiently combines genetic perturbations with mRNA barcodes for advanced pooled screening applications.

Keywords:
CRISPR Cas9CRISPR knockoutCRISPRiPerturb-seqgenetic interactionshigh-content screeningmultiplexed perturbationoptical pooled screens

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Last Updated: Jun 16, 2026

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • Forward genetic screens are crucial for understanding biological systems by analyzing gene perturbations and resulting phenotypes.
  • Multiplexing genetic perturbations enhances screening efficiency and enables the study of complex genetic interactions.
  • Existing multiplexing tools often conflict with pooled screening methods requiring mRNA-embedded barcodes, limiting their use in techniques like single-cell sequencing.

Purpose of the Study:

  • To develop a novel lentiviral system, CROPseq-multi, for multiplexing Streptococcus pyogenes (Sp) Cas9-based perturbations with mRNA-embedded barcodes.
  • To ensure compatibility with pooled screening methodologies, including those using single-cell sequencing.
  • To improve the efficiency and applicability of multiplexed genetic screens.

Main Methods:

  • Development of a CROPseq-inspired lentiviral system (CROPseq-multi).
  • Integration of Streptococcus pyogenes (Sp) Cas9 for genetic perturbations.
  • Incorporation of mRNA-embedded barcodes for pooled screening compatibility.
  • Optimization of an in situ detection protocol for optical pooled screens.

Main Results:

  • CROPseq-multi demonstrates equivalent per-guide activity to CROPseq with low lentiviral recombination.
  • The system is compatible with enrichment screening and optical pooled screens.
  • Optimized protocols improved barcode detection efficiency 10-fold and decoding efficiency 3-fold for optical pooled screens.
  • CROPseq-multi is extensible to single-cell sequencing readouts.

Conclusions:

  • CROPseq-multi provides a versatile solution for multiplexed genetic screening using SpCas9.
  • This advancement broadens the applicability of pooled screening methodologies, particularly those requiring mRNA-embedded barcodes.
  • Facilitates deeper investigation into genetic interactions and complex biological systems.