ArticleScientific reports2017
CRISPR/Cas9-mediated gene manipulation to create single-amino-acid-substituted and floxed mice with a cloning-free method.
Article in Scientific reports, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 30 papers.
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Who cites it
30 citing papers in PubMed, 54 citations in OpenAlex.
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- SALL1 enforces microglia-specific DNA binding and function of SMADs to establish microglia identity.Nature immunology · 2023Article
- Research and Therapeutic Approaches in Stem Cell Genome Editing by CRISPR Toolkit.Molecules (Basel, Switzerland) · 2023Review
- CRISPR/Cas9-generated mouse model with humanizing single-base substitution in theFrontiers in genome editing · 2023Article
- A dominant function of LynB kinase in preventing autoimmunity.Science advances · 2022Article
- Highly reliable creation of floxed alleles by electroporating single-cell embryos.BMC biology · 2022Article
- Understanding the molecular basis of cardiomyopathy.American journal of physiology. Heart and circulatory physiology · 2022Review
- Desmosomal COP9 regulates proteome degradation in arrhythmogenic right ventricular dysplasia/cardiomyopathy.The Journal of clinical investigation · 2021Article
- Response to "Reproducibility of CRISPR-Cas9 methods for generation of conditional mouse alleles: a multi-center evaluation".Genome biology · 2021Article
- Homozygous G650del nexilin variant causes cardiomyopathy in mice.JCI insight · 2020Article
- Atypical ALPK2 kinase is not essential for cardiac development and function.American journal of physiology. Heart and circulatory physiology · 2020Article
- Hyperglycemia Acutely Increases Cytosolic Reactive Oxygen Species viaCirculation research · 2020Article
- Article
- A Simple Cloning-free Method to Efficiently Induce Gene Expression Using CRISPR/Cas9.Molecular therapy. Nucleic acids · 2019Article
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Authors and funding
7 authors at 2 institutions in 2 countries.
Funding
Abstract
Clustered regulatory interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) technology is a powerful tool to manipulate the genome with extraordinary simplicity and speed. To generate genetically modified animals, CRISPR/Cas9-mediated genome editing is typically accomplished by microinjection of a mixture of Cas9 DNA/mRNA and single-guide RNA (sgRNA) into zygotes. However, sgRNAs used for this approach require manipulation via molecular cloning as well as in vitro transcription. Beyond these complexities, most mutants obtained with this traditional approach are genetically mosaic, yielding several types of cells with different genetic mutations. Recently, a growing body of studies has utilized commercially available Cas9 protein together with sgRNA and a targeting construct to introduce desired mutations. Here, we report a cloning-free method to target the mouse genome by pronuclear injection of a commercial Cas9 protein:crRNA:tracrRNA:single-strand oligodeoxynucleotide (ssODN) complex into mouse zygotes. As illustration of this method, we report the successful generation of global gene-knockout, single-amino-acid-substituted, as well as floxed mice that can be used for conditional gene-targeting. These models were produced with high efficiency to generate non-mosaic mutant mice with a high germline transmission rate.
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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.