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Knocking in Multifunctional Gene Tags into SMC Complex Subunits Using Gene Editing.

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|June 1, 2019
PubMed
Summary

Researchers can now precisely study the condensin complex in vertebrates using CRISPR/Cas9 gene editing. This technique allows for detailed analysis of condensin

Keywords:
CRISPR/Cas9ChromosomesCondensinGene editingMitosisSouthern blot hybridizationStreptavidin binding peptide

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Area of Science:

  • Genetics and Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Condensin is a crucial pentameric complex for genome organization and folding in vertebrates.
  • Studying endogenous condensin function requires precise genetic manipulation tools.
  • Previous methods lacked the precision to analyze condensin in its native cellular environment.

Purpose of the Study:

  • To detail methods for engineering vertebrate cells to study the condensin complex.
  • To enable precise biochemical and microscopic analysis of endogenous condensin.
  • To facilitate a deeper understanding of condensin's role in genome organization.

Main Methods:

  • Utilizing CRISPR/Cas9 gene editing to knock protein tags into endogenous loci.
  • Employing streptavidin binding peptide (SBP) tags for proteomic applications.
  • Engineering cells for affinity, fluorescent, or multifunctional tagging of condensin.

Main Results:

  • Established the first genome-wide map of condensin using SBP tagging.
  • Enabled detailed biochemical and microscopic analysis of tagged condensin.
  • Provided researchers with tools for high-precision study of condensin in vivo.

Conclusions:

  • CRISPR/Cas9 facilitates precise engineering of cells for studying protein complexes like condensin.
  • Tagged endogenous condensin allows for comprehensive analysis of its functions.
  • This approach offers unprecedented precision for understanding condensin's cellular behavior and interactions.