무세포 시스템 기반 유전자 발현 모형 개발에 대한 연구

다국어 초록 (Multilingual Abstract)

In synthetic biology, the design of genetic circuits commonly involves selecting genetic elements such as promoters, ribosome binding sites, and terminators. However, selecting an appropriate combination that ensures the circuit function as designed becomes increasingly challenging as the number of genetic parts grows. This study introduces a novel modeling approach, explaining gene expression through the incorporation of reaction equations for transcription and translation processes. Our model applies quantitative characteristics of genetic parts to these reaction equations and is constructed using a cell-free system to minimize biological uncertainties. Parameter estimation was executed using the Hamiltonian monte carlo sampling method, followed by simulations via ordinary differential equation solvers to validate the model against real data. We initially estimated unknown parameters with only transcription equations under three distinct conditions. Subsequently, the parameters in translation reactions were inferred using the three variant sets of transcription parameters. The model can be extended to the simulations involving multiple genes and living cells. We expect this modeling approach enables the prediction and evaluation of circuit designs without necessitating experiments, thereby significantly curtailing time and labor in synthetic biology research.

목차 (Table of Contents)

• 충 남 대 학 교 대 학 원
• 목 차
• 목차 ⅰ
• 약어 ⅱ
• 표 목차 ⅲ
• 그림 목차 ⅳ
• I. 서론 12
• 1.1 합성생물학 12
• 1.1.1. 생물학적 부품(BioBrick) 14
• 1.1.2. 유전자 회로 15
• 1.2 무세포 단백질 합성 시스템 17
• 1.2.1 센트럴 도그마(Central dogma) 20
• 1.2.2 PURE system 23
• 1.3 수학적 모형과 시뮬레이션 24
• 1.4 본 연구의 목적 26
• II. 재료 및 실험 방법 29
• 2.1. 발현 유전자 29
• 2.2. 단백질 정량 30
• 2.2.1 세포 배양 및 회수 30
• 2.2.2 FPLC 단백질 정제 32
• 2.2.3 브래드퍼드 단백질 정량법 35
• 2.3. 무세포 단백질 발현 및 측정 36
• 2.4. 수학적 모델링 37
• 2.4.1 발현 모델 정의 37
• 2.4.2 상미분 방정식 solver 47
• 2.4.3 Hamiltonian Monte Carlo 49
• Ⅲ. 결과 및 고찰 53
• 3.1 전사 모형 검증 53
• 3.2 Standard Curve 작성 57
• 3.3 전사 번역 모형 검증 59
• 3.4 바이오 부품 최적화 및 단백질 발현 극대화 73
• 3.5 어플리케이션 77
• Ⅳ. 결론 79
• Ⅴ. 참고문헌 81
• ABSTRACT 87