CRISPR/Cas9-mediated genome editing in naïve human embryonic stem cells

Sci Rep. 2017 Nov 30;7(1):16650. doi: 10.1038/s41598-017-16932-y.


The combination of genome-edited human embryonic stem cells (hESCs) and subsequent neural differentiation is a powerful tool to study neurodevelopmental disorders. Since the naïve state of pluripotency has favourable characteristics for efficient genome-editing, we optimized a workflow for the CRISPR/Cas9 system in these naïve stem cells. Editing efficiencies of respectively 1.3-8.4% and 3.8-19% were generated with the Cas9 nuclease and the D10A Cas9 nickase mutant. Next to this, wildtype and genome-edited naïve hESCs were successfully differentiated to neural progenitor cells. As a proof-of-principle of our workflow, two monoclonal genome-edited naïve hESCs colonies were obtained for TUNA, a long non-coding RNA involved in pluripotency and neural differentiation. In these genome-edited hESCs, an effect was seen on expression of TUNA, although not on neural differentiation potential. In conclusion, we optimized a genome-editing workflow in naïve hESCs that can be used to study candidate genes involved in neural differentiation and/or functioning.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • CRISPR-Cas Systems*
  • Cell Differentiation / genetics
  • Cell Line
  • Clustered Regularly Interspaced Short Palindromic Repeats
  • Gene Editing*
  • High-Throughput Nucleotide Sequencing
  • Human Embryonic Stem Cells / metabolism*
  • Humans
  • Neural Stem Cells / cytology
  • Neural Stem Cells / metabolism
  • Workflow