ArticleMethods in molecular biology (Clifton, N.J.)2022
Gene Editing in Human Induced Pluripotent Stem Cells Using Doxycycline-Inducible CRISPR-Cas9 System.
Article in Methods in molecular biology (Clifton, N.J.), 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 4 papers.
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Who cites it
4 citing papers in PubMed, 10 citations in OpenAlex.
- CRISPR/Cas9 and iPSC-Based Therapeutic Approaches in Alzheimer's Disease.Antioxidants (Basel, Switzerland) · 2025Review
- Efficient gene editing in induced pluripotent stem cells enabled by an inducible adenine base editor with tunable expression.Scientific reports · 2023Article
- Efficient deletion of microRNAs using CRISPR/Cas9 with dual guide RNAs.Frontiers in molecular biosciences · 2023Article
- Principles and Applications of CRISPR Toolkit in Virus Manipulation, Diagnosis, and Virus-Host Interactions.Cells · 2022Review
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Authors and funding
3 authors at 2 institutions in 1 country.
Funding
Abstract
Induced pluripotent stem cells (iPSCs) generated from patients are a valuable tool for disease modelling, drug screening, and studying the functions of cell/tissue-specific genes. However, for this research, isogenic iPSC lines are important for comparison of phenotypes in the wild type and mutant differentiated cells generated from the iPSCs. The advent of gene editing technologies to correct or generate mutations helps in the generation of isogenic iPSC lines with the same genetic background. Due to the ease of programming, CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-Cas9-based gene editing tools have gained pace in gene manipulation studies, including investigating complex diseases like cancer. An iPSC line with drug inducible Cas9 expression from the Adeno-Associated Virus Integration Site 1 (AAVS1) safe harbor locus offers a controllable expression of Cas9 with robust gene editing. Here, we describe a stepwise protocol for the generation and characterization of such an iPSC line (AAVS1-PDi-Cas9 iPSC) with a doxycycline (dox)-inducible Cas9 expression cassette from the AAVS1 safe harbor site and efficient editing of target genes with lentiviral vectors expressing gRNAs. This approach with a tunable Cas9 expression that allows investigating gene functions in iPSCs or in the differentiated cells can serve as a versatile tool in disease modelling studies.
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Registered trials
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