A Portable Molecular Diagnostic System Integrating On-Chip Gene Amplification and Non-Spectroscopic Signal Analysis
- 주제(키워드) Molecular diagnostic , Loop-mediated isothermal amplification , Retroreflective Janus particle , Salmonella Typhimurium
- 주제(DDC) 547
- 발행기관 아주대학교 일반대학원
- 지도교수 윤현철
- 발행년도 2026
- 학위수여년월 2026. 2
- 학위명 석사
- 학과 및 전공 일반대학원 분자과학기술학과
- 실제URI http://www.dcollection.net/handler/ajou/000000035625
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
Polymerase Chain Reaction (PCR) is a widely used molecular diagnostic method but requires expensive and complex equipment to detect low concentrations of target DNA. To address this limitation, we have developed a compact and cost-effective molecular diagnosis platform by integrating loop-mediated isothermal amplification (LAMP) with a simple optical detection system. To minimize the DNA amplification steps, we employed the loop- mediated isothermal amplification (LAMP) method to enable DNA amplification on a small Peltier device. This constant-temperature device can replace bulky thermocyclers when combined with the LAMP method. For detection, we introduced Retroreflective Janus Particles (RJPs) as non-spectroscopic optical probes to generate signals using a white LED and a camera. The system uses specially designed primers that produce double-stranded amplicons with single-stranded DNA (ssDNA) and biotin at each 5' end, forming a sandwich-type complex with a DNA-immobilized substrate and an avidin-functionalized optical probe. In this study, we developed a platform that integrates these processes into a single chip. Using this platform, we successfully quantified the foodborne pathogen Salmonella by capturing and counting the optical probes with simple imaging techniques. Our approach shows that molecular diagnostics can be performed with portable cost-effective devices, providing a practical solution for point-of-care testing and early detection of foodborne pathogens.
more목차
1. Introduction 1
1.1 Limitations of the conventional molecular diagnostic method 1
1.2 Isothermal amplification for portable molecular diagnostic platform 2
1.3 Retroreflective Janus particles for non-spectroscopic detection 5
1.4 Integrated diagnostic platform for point-of-care applications 6
1.5 Aim of the thesis 10
2. Materials and methods 12
2.1 Reagents and apparatus 12
2.2 Fabrication of the molecular diagnostic chip 14
2.2.1 Modification of complementary ssDNA on sensing surface 14
2.2.2 Fabrication of sensing chip with PDMS 16
2.3 Amplification of target gene 17
2.3.1 Primer design for LAMP assay 17
2.3.2 Preparation of target genes for LAMP assay 19
2.3.3 LAMP assay with amplification module 19
2.4 Purification of amplified products 21
2.4.1 Purification of Amplified Products with gel electrophoresis module 21
2.4.2 Optimization of Purification Conditions 21
2.4.3 Feasibility Test of the Purification Module 22
2.5 Detection of Salmonella using the developed integrated system 24
2.5.1 Fabrication of avidin-conjugated RJPs 24
2.5.2 Quantitative analysis of Salmonella 24
2.5.3 Comparison of analytical performance with real-time LAMP 25
3. Results and discussion 26
3.1 Confirmation of ssDNA modification on sensing surface 26
3.2 Amplification of target gene 29
3.2.1 Construction of amplification module with Peltier element 29
3.2.2 Optimization of LAMP method on amplification module 32
3.3 Purification of amplified LAMP products 34
3.3.1 Construction of miniaturized gel electrophoresis purification module 34
3.3.2 Optimization of purification conditions 36
3.3.3 Feasibility test of developed purification module 40
3.4 Detection of Salmonella using the developed integrated system 43
3.4.1 Verification of optical signaling probe 43
3.4.2 Optimization of RJP reaction time 45
3.4.3 Quantitative analysis of Salmonella 47
3.4.4 Comparison of analytical performance with real-time LAMP 50
4. Conclusions 52
5. References 54
