ArticleScientific reports2015
CRISPR/Cas9-mediated reporter knock-in in mouse haploid embryonic stem cells.
Article in Scientific reports, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 14 papers.
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Who cites it
14 citing papers in PubMed, 32 citations in OpenAlex.
- Efficient tagging of endogenous proteins in human cell lines for structural studies by single-particle cryo-EM.Proceedings of the National Academy of Sciences of the United States of America · 2023Article
- EditingInternational journal of molecular sciences · 2021Review
- Gene insertion in3 Biotech · 2021Article
- Efficient SNP editing in haploid human pluripotent stem cells.Journal of assisted reproduction and genetics · 2020Article
- Human Genomic Safe Harbors and the Suicide Gene-Based Safeguard System for iPSC-Based Cell Therapy.Stem cells translational medicine · 2019Article
- Unbiased compound-protein interface mapping and prediction of chemoresistance loci through forward genetics in haploid stem cells.Oncotarget · 2018Article
- A versatile genetic tool: haploid cells.Stem cell research & therapy · 2017Review
- Targeted Genome Replacement via Homology-directed Repair in Non-dividing Cardiomyocytes.Scientific reports · 2017Article
- CRISPR/Cas9: Transcending the Reality of Genome Editing.Molecular therapy. Nucleic acids · 2017Review
- Phenotypic screening with primary neurons to identify drug targets for regeneration and degeneration.Molecular and cellular neurosciences · 2017Review
- Generation of genetically modified mice using CRISPR/Cas9 and haploid embryonic stem cell systems.Dong wu xue yan jiu = Zoological research · 2016Review
- Efficient generation of FVII gene knockout mice using CRISPR/Cas9 nuclease and truncated guided RNAs.Scientific reports · 2016Article
- CRISPR-Cas9 directed knock-out of a constitutively expressed gene using lance array nanoinjection.SpringerPlus · 2016Article
- Genome Editing Using Mammalian Haploid Cells.International journal of molecular sciences · 2015Review
Corrections and comments
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Authors and funding
8 authors at 5 institutions in 4 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Mouse parthenogenetic haploid embryonic stem cells (ESCs) are pluripotent cells generated from chemically activated oocytes. Haploid ESCs provide an opportunity to study the effect of genetic alterations because of their hemizygotic characteristics. However, their further application for the selection of unique phenotypes remains limited since ideal reporters to monitor biological processes such as cell differentiation are missing. Here, we report the application of CRISPR/Cas9-mediated knock-in of a reporter cassette, which does not disrupt endogenous target genes in mouse haploid ESCs. We first validated the system by inserting the P2A-Venus reporter cassette into the housekeeping gene locus. In addition to the conventional strategy using the Cas9 nuclease, we employed the Cas9 nickase and truncated sgRNAs to reduce off-target mutagenesis. These strategies induce targeted insertions with an efficiency that correlated with sgRNA guiding activity. We also engineered the neural marker gene Sox1 locus and verified the precise insertion of the P2A-Venus reporter cassette and its functionality by monitoring neural differentiation. Our data demonstrate the successful application of the CRISPR/Cas9-mediated knock-in system for establishing haploid knock-in ESC lines carrying gene specific reporters. Genetically modified haploid ESCs have potential for applications in forward genetic screening of developmental pathways.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.