Human pluripotent stem cells (hPSCs) offer powerful platforms for studying mechanisms of human diseases and for evaluating potential treatments. Genome editing, particularly the CRISPR/Cas9-based method, is highly effective for generating cell and animal models to study genetic human diseases. However, the procedure for generating gene-edited hPSCs is laborious, time consuming and unintentional genetic changes may confound the consequent experiments and conclusions. Here we describe one-step knockin of the NanoLuc luciferase gene (Nluc) to the fragile X syndrome gene, FMR1, in a human embryonic stem cell line (hESC), H1, and a fragile X disease model human induced pluripotent stem cell line (hiPSC), FX-iPSC. The luciferase reporter cell lines provide new platforms for exploring potential treatments for fragile X syndrome. The shortened and scarless targeting method described here can be effectively applied to other genes.
CITATION STYLE
Li, M., Hunt, J. F. V. S., Bhattacharyya, A., & Zhao, X. (2019). One-step generation of seamless luciferase gene knockin using CRISPR/Cas9 genome editing in human pluripotent stem cells. In Methods in Molecular Biology (Vol. 1942, pp. 61–69). Humana Press Inc. https://doi.org/10.1007/978-1-4939-9080-1_5
Mendeley helps you to discover research relevant for your work.