論文 - 詳細
| RRC ID | 77239 |
|---|---|
| 著者 | Zhang H, Zhou Q, Chen H, Lu D. |
| タイトル | Prime Editor 3 Mediated Beta-Thalassemia Mutations of the HBB Gene in Human Erythroid Progenitor Cells. |
| ジャーナル | Int J Mol Sci |
| Abstract |
Recently developed Prime Editor 3 (PE3) has been implemented to induce genome editing in various cell types but has not been proven in human hematopoietic stem and progenitor cells. Using PE3, we successfully installed the beta-thalassemia (beta-thal) mutations in the HBB gene in the erythroid progenitor cell line HUDEP-2. We inserted the mCherry reporter gene cassette into editing plasmids, each including the prime editing guide RNA (pegRNA) and nick sgRNA. The plasmids were electroporated into HUDEP-2 cells, and the PE3 modified cells were identified by mCherry expression and collected using fluorescence-activated cell sorting (FACS). Sanger sequencing of the positive cells confirmed that PE3 induced precise beta-thal mutations with editing ratios from 4.55 to 100%. Furthermore, an off-target analysis showed no unintentional edits occurred in the cells. The editing ratios and parameters of pegRNA and nick sgRNA were also analyzed and summarized and will contribute to enhanced PE3 design in future studies. The characterization of the HUDEP-2 beta-thal cells showed typical thalassemia phenotypes, involving ineffective erythropoiesis, abnormal erythroid differentiation, high apoptosis rate, defective alpha-globin colocalization, cell viability deterioration, and ROS resisting deficiency. These HUDEP-2 beta-thal cells could provide ideal models for future beta-thal gene therapy studies. |
| 巻・号 | 23(9) |
| 公開日 | 2022-4-30 |
| DOI | 10.3390/ijms23095002 |
| PII | ijms23095002 |
| PMID | 35563395 |
| PMC | PMC9099916 |
| MeSH | Erythroid Precursor Cells* / metabolism Gene Editing* / methods Hemoglobins* / genetics Hemoglobins* / therapeutic use Humans Mutation alpha-Globins / genetics beta-Globins / genetics beta-Thalassemia* / genetics beta-Thalassemia* / therapy |
| IF | 4.556 |
| オルトメトリクス指標 |
オルトメトリクス指標項目
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| 最多言及媒体 | X(Twitter) |
| 各媒体での言及数の合計 | 1 |
| 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 | |
| ヒト・動物細胞 | HUDEP-2(RCB4557) |