Development of a Synthetic UPR Monitoring System with Signal Amplification in Chinese Hamster Ovary Cells
- 주제(키워드) ER stress , Unfolded protein response , Synthetic biology , Signal amplification , CHO cells
- 주제(DDC) 547
- 발행기관 아주대학교 일반대학원
- 지도교수 Pyung Cheon Lee
- 발행년도 2026
- 학위수여년월 2026. 2
- 학위명 석사
- 학과 및 전공 일반대학원 분자과학기술학과
- 실제URI http://www.dcollection.net/handler/ajou/000000035750
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
목차
1. Introduction 1
2. Materials and Methods 4
2.1. Plasmid construction 4
2.2. Cell culture and maintenance 4
2.3. Cell transfection 5
2.4. Generation of stable cell pools harboring a UPR-dependent genetic circuit via recombinase-mediated cassette exchange (RMCE) 5
2.5. Chemical treatment 6
2.6. Fluorescence expression analysis using flow cytometry 7
2.7. Preparation of RNA and cDNA 8
2.8 Evaluation of relative mRNA expression level using qRT-PCR 8
2.9. Regression model 9
2.10. Statistical analysis 9
3. Results 25
3.1. A UPR-responsive synthetic promoter exhibits low inducibility under ER stress 25
3.2. Transcriptional cascade and positive feedback enable UPR- dependent signal amplification under ER stress 27
3.3. Stable expression of a transcriptional cascade with positive feedback enables UPR-dependent signal amplification 32
3.4. Real-time monitoring of UPR dynamics is limited by the long half- life of EGFP and positive feedback-driven maintenance of Tet-On3G after stress relief 37
4. Discussion 41
5. References 45
