論文 - 詳細
| RRC ID | 73354 |
|---|---|
| 著者 | Ishibashi R, Maki R, Kitano S, Miyachi H, Toyoshima F. |
| タイトル | Development of an in vivo cleavable donor plasmid for targeted transgene integration by CRISPR-Cas9 and CRISPR-Cas12a. |
| ジャーナル | Sci Rep |
| Abstract |
The CRISPR-Cas system is widely used for genome editing of cultured cells and organisms. The discovery of a new single RNA-guided endonuclease, CRISPR-Cas12a, in addition to the conventional CRISPR-Cas9 has broadened the number of editable target sites on the genome. Here, we developed an in vivo cleavable donor plasmid for precise targeted knock-in of external DNA by both Cas9 and Cas12a. This plasmid, named pCriMGET_9-12a (plasmid of synthetic CRISPR-coded RNA target sequence-equipped donor plasmid-mediated gene targeting via Cas9 and Cas12a), comprises the protospacer-adjacent motif sequences of Cas9 and Cas12a at the side of an off-target free synthetic CRISPR-coded RNA target sequence and a multiple cloning site for donor cassette insertion. pCriMGET_9-12a generates a linearized donor cassette in vivo by both CRISPR-Cas9 and CRISPR-Cas12a, which resulted in increased knock-in efficiency in culture cells. This method also achieved > 25% targeted knock-in of long external DNA (> 4 kb) in mice by both CRISPR-Cas9 and CRISPR-Cas12a. The pCriMGET_9-12a system expands the genomic target space for transgene knock-in and provides a versatile, low-cost, and high-performance CRISPR genome editing tool. |
| 巻・号 | 12(1) |
| ページ | 17775 |
| 公開日 | 2022-10-22 |
| DOI | 10.1038/s41598-022-22639-6 |
| PII | 10.1038/s41598-022-22639-6 |
| PMID | 36272994 |
| PMC | PMC9588054 |
| MeSH | Animals CRISPR-Cas Systems* DNA Endonucleases / genetics Gene Editing* / methods Mice Plasmids / genetics RNA / genetics Transgenes |
| IF | 3.998 |
| オルトメトリクス指標 |
オルトメトリクス指標項目
|
| 最多言及媒体 | X(Twitter) |
| 各媒体での言及数の合計 | 4 |
| 過去6か月間でのオルトメトリクス指標の変動値 | 0.0 |
| リソース情報 | |
| 遺伝子材料 | CSII-EF-mRFP1-RfA (RDB012128) pCriMGET_9-12a (RDB20013) |
| ヒト・動物細胞 | 293T(RCB2202) |