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Highly Efficient Mouse Genome Editing by CRISPR Ribonucleoprotein Electroporation of Zygotes
| Content Provider | Scilit |
|---|---|
| Author | Chen, Sean Lee, Benjamin Lee, Angus Yiu-Fai Modzelewski, Andrew J. He, Lin |
| Copyright Year | 2016 |
| Description | Journal: Journal of Biological Chemistry The CRISPR/Cas9 system has been employed to efficiently edit the genomes of diverse model organisms. CRISPR-mediated mouse genome editing is typically accomplished by microinjection of Cas9 DNA/RNA and single guide RNA (sgRNA) into zygotes to generate modified animals in one step. However, microinjection is a technically demanding, labor-intensive, and costly procedure with poor embryo viability. Here, we describe a simple and economic electroporation-based strategy to deliver Cas9/sgRNA ribonucleoproteins into mouse zygotes with 100% efficiency for in vivo genome editing. Our methodology, designated as CRISPR RNP Electroporation of Zygotes (CRISPR-EZ), enables highly efficient and high-throughput genome editing in vivo, with a significant improvement in embryo viability compared with microinjection. Using CRISPR-EZ, we generated a variety of editing schemes in mouse embryos, including indel (insertion/deletion) mutations, point mutations, large deletions, and small insertions. In a proof-of-principle experiment, we used CRISPR-EZ to target the tyrosinase (Tyr) gene, achieving 88% bi-allelic editing and 42% homology-directed repair-mediated precise sequence modification in live mice. Taken together, CRISPR-EZ is simple, economic, high throughput, and highly efficient with the potential to replace microinjection for in vivo genome editing in mice and possibly in other mammals. |
| Related Links | http://www.jbc.org/content/291/28/14457.full.pdf https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4938170/pdf http://www.jbc.org/article/S0021925820349073/pdf |
| Ending Page | 14467 |
| Page Count | 11 |
| Starting Page | 14457 |
| ISSN | 00219258 |
| e-ISSN | 1083351X |
| DOI | 10.1074/jbc.m116.733154 |
| Journal | Journal of Biological Chemistry |
| Issue Number | 28 |
| Volume Number | 291 |
| Language | English |
| Publisher | Elsevier BV |
| Publisher Date | 2016-07-01 |
| Access Restriction | Open |
| Subject Keyword | Journal: Journal of Biological Chemistry Developmental Biology Crispr, Cas9, Genome Editing, Electroporation, Rnp Gene Knockout Ribonuclear Protein (rnp) |
| Content Type | Text |
| Resource Type | Article |
| Subject | Cell Biology Biochemistry Molecular Biology |
