Zinc Finger Nuclease-Based Genome Editing: A Technique for Modifying Genome in Human Pluripotent Stem Cells by Double-Stranded Homology Dependant Repair Mechanism
This video demonstrates a technique of zinc-finger nuclease-based genome editing in human pluripotent stem cells (hPSCs). As hPSCs can potentially be differentiated into various cell types, this technology is helpful for in vitro pathological and pharmacological studies in a patient-specific manner.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Prepare Stem Cells for Editing Grow and culture human pluripotent stem cells (hPSCs) on a 6-well plate containing 2.4 × 106 cells/plate of mitomycin C-inactivated mouse embryonic fibroblast (MEF) feeders grown on gelatin. Maintain hPSCs in 3 ml of human embryonic stem cell media per well (hESC media…
Representative Results
Figure 1. Schematic of gene edited AAVS1 locus using the AAV-CAGGS-EGFP repair template. Modified from Hockemeyer et al., 2009.
Zinc Finger Nuclease-Based Genome Editing: A Technique for Modifying Genome in Human Pluripotent Stem Cells by Double-Stranded Homology Dependant Repair Mechanism. J. Vis. Exp. (Pending Publication), e20972, doi: (2023).