Experiment: Testing AAV5 for activation of tdTomato in mouse airway club and ciliated cells
PI: Guang-Ping Gao, PhD
Description: AAV2/5 mediated gene editing in the mouse airway was tested by deliverying SpCas9 and guide RNAs targeting the Ai9 transgene in Ai9 transgenic mice. Gene editing quantified by tdTomato activation and cell specific markers for club and ciliated cell types.
Editing Assay:
Editing efficiency calculated by counting cells double positive for tdTomato and CCSP (a marker of club/clara cells) and acetylated tubulin (a marker of ciliated cells) compared to DAPI stained cells. Analyzed 5 sections per mouse.
Parent Project: Develop Combinatorial Non-Viral and Viral CRISPR Delivery for Lung Diseases
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Other experiments in this project: 2
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Results |
| Editing Efficiency | ||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Condition | Tissue | Cell Type | Sex | Editor | Model | Target Locus | Guide | Vector | Dosage | Injection Frequency | % cells | Image |
| AAV-SpCas9 | epithelium of bronchus (TARGET) | club cell | Female | SpCas9 | Ai9 mouse | AAV.pU1a-SpCas9 | 6e10 gc/each AAV per mouse by intratracheal injection | once | ||||
| AAV-SpCas9 | epithelium of bronchus (TARGET) | ciliated cell of the bronchus | Female | SpCas9 | Ai9 mouse | AAV.pU1a-SpCas9 | 6e10 gc/each AAV per mouse by intratracheal injection | once | ||||
| Dual AAV | epithelium of bronchus (TARGET) | club cell | Female | SpCas9 | Ai9 mouse | Ai9/Ai14 reporter transgene | sgAi9R; sgAi9L | AAV.pU1a-SpCas9; H509 AAVsc-u6-sgAI9L-U6-AI9R-U1A-EGFP (1) | 6e10 gc/each AAV per mouse by intratracheal injection | once | ||
| Dual AAV | epithelium of bronchus (TARGET) | ciliated cell of the bronchus | Female | SpCas9 | Ai9 mouse | Ai9/Ai14 reporter transgene | sgAi9R; sgAi9L | AAV.pU1a-SpCas9; H509 AAVsc-u6-sgAI9L-U6-AI9R-U1A-EGFP (1) | 6e10 gc/each AAV per mouse by intratracheal injection | once | ||
| AAV5 mediates genome editing in club and ciliated cells. |
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