| RRC ID |
77294
|
| 著者 |
Suzuki M, Iida M, Hayashi T, Suzuki KT.
|
| タイトル |
CRISPR-Cas9-Based Functional Analysis in Amphibians: Xenopus laevis, Xenopus tropicalis, and Pleurodeles waltl.
|
| ジャーナル |
Methods Mol Biol
|
| Abstract |
Amphibians have made many fundamental contributions to our knowledge, from basic biology to biomedical research on human diseases. Current genome editing tools based on the CRISPR-Cas system enable us to perform gene functional analysis in vivo, even in non-model organisms. We introduce here a highly efficient and easy protocol for gene knockout, which can be used in three different amphibians seamlessly: Xenopus laevis, Xenopus tropicalis, and Pleurodeles waltl. As it utilizes Cas9 ribonucleoprotein complex (RNP) for injection, this cloning-free method enables researchers to obtain founder embryos with a nearly complete knockout phenotype within a week. To evaluate somatic mutation rate and its correlation to the phenotype of a Cas9 RNP-injected embryo (crispant), we also present accurate and cost-effective genotyping methods using pooled amplicon-sequencing and a user-friendly web-based tool.
|
| 巻・号 |
2637
|
| ページ |
341-357
|
| 公開日 |
2023-1-1
|
| DOI |
10.1007/978-1-0716-3016-7_26
|
| PMID |
36773159
|
| MeSH |
Animals
CRISPR-Cas Systems* / genetics
Gene Editing / methods
Humans
Pleurodeles* / genetics
Xenopus / genetics
Xenopus laevis / genetics
|
| リソース情報 |
| ツメガエル・イモリ |
Xenopus tropicalis
Pleurodeles waltl |
