Small molecules enhance CRISPR genome editing in pluripotent stem cells

Cell Stem Cell. 2015 Feb 5;16(2):142-7. doi: 10.1016/j.stem.2015.01.003.

Abstract

The bacterial CRISPR-Cas9 system has emerged as an effective tool for sequence-specific gene knockout through non-homologous end joining (NHEJ), but it remains inefficient for precise editing of genome sequences. Here we develop a reporter-based screening approach for high-throughput identification of chemical compounds that can modulate precise genome editing through homology-directed repair (HDR). Using our screening method, we have identified small molecules that can enhance CRISPR-mediated HDR efficiency, 3-fold for large fragment insertions and 9-fold for point mutations. Interestingly, we have also observed that a small molecule that inhibits HDR can enhance frame shift insertion and deletion (indel) mutations mediated by NHEJ. The identified small molecules function robustly in diverse cell types with minimal toxicity. The use of small molecules provides a simple and effective strategy to enhance precise genome engineering applications and facilitates the study of DNA repair mechanisms in mammalian cells.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Clustered Regularly Interspaced Short Palindromic Repeats / genetics*
  • DNA Repair / drug effects*
  • Dose-Response Relationship, Drug
  • Genetic Engineering / methods*
  • Genome, Human / genetics*
  • HeLa Cells
  • High-Throughput Screening Assays
  • Humans
  • Molecular Structure
  • Pluripotent Stem Cells / drug effects*
  • Pluripotent Stem Cells / metabolism*
  • Small Molecule Libraries / chemistry
  • Small Molecule Libraries / pharmacology*
  • Structure-Activity Relationship

Substances

  • Small Molecule Libraries