신속한 DNA정량을 위한 Labchip 기반의 실시간 PCR법 개발
Development of Labchip Based Realtime PCR for Rapid Human DNA Quantification
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초록
법유전학에서 DNA 정량 과정은 정확한 DNA형을 얻기 위한 필수과정이다. 기존 실시간 증폭시스템을 이용한 정량 분석법은 시약의 높은 단가와 오랜 시간소요 등과 같은 단점을 가지고 있다. 본연구에서는 샘플 내 존재하는 특정 인간 DNA양을 측정할 수 있는 신속 DNA 정량 시험법을 개발하여 기존 시스템의 단점을 보완하였다. 개발된 시험법은 실시간 중합효소 연쇄반응(Realtime PCR) 기술을 기반으로 Microfluidic chip (Lab-on-a-chip, LOC)을 이용하여 정확한 분석 값을 기존 대비 5배 단축된 시간(15분) 안에 얻을 수 있었다. 또한, 표준 DNA를 사용하여 시험방법의 정확성, 재현성, 민감성및 종 특이성을 검사하였고 현장 증거물에 대한 적용 가능성 및 상용 키트에 포함된 정량 시약과의 호환성을 테스트하여 적용 범위의 다양성을 확인하였다.
Abstract
Human DNA quantification on DNA profiling is an important process to get clear genotypes without any allele drop-out and peak imbalances in forensic genetics. Conventional realtime PCR for human DNA quantification is a time-consuming and expensive technique. In order to overcome these disadvantages we developed UltraFast Realtime PCR system based on the Microfluidic chip(Lab-on-a-chip, LOC) technology. This system can quickly and precisely measure DNA quantification within 15 minutes which was reducing 5 times than conventional method. We validated the new system based on standard DNA using various measures, such as accuracy, reproducibility, sensitivity, and human specificity. We confirmed it's applicability to the cases of crime scene and it's compatibility with other quantification kits.
참고문헌 (18)
- RL Green, "Development validation of the Quantifiler TM real-time PCR kits for the quantification of human nuclear DNA samples", J Forensic Sci, 50(4), pp.809~825 (2005)
- JA Niclas, "Development of an Alubased, real-time PCR method of quantitation of human DNA in forensic sample", J Forensic Sci, 48(5), pp.936~944 (2003)
- 박정현, "실시간 중합효소연쇄반응을 이용한 유전자 정량분석법의 시험방법 유효화", 과학수사학회지, 5(4), pp.336~343 (2011), 한국과학수사학회
- S Palmer, "New real-time reverse transcriptase-initiated PCR assay with single-copy sensitivity for human immunodeficiency virus type 1 RNA in plasma", J Clin Micobiol, 41, pp.4531~4536 (2003)
- IG Robert, "Development of a Real-Time qPCR Assay for Quantification of Covert Baculovirus Infections in a Major African Crop Pest", Insects, 6, pp.746~759 (2015)
- V Fajardo, "Real-Time PCR for detection and quantification of red deer (Cervus elaphus), fallow deer (Dama dama), and roe deer (Capreolus capreolus) in meat mixtures", Meat Sci, 78(2), pp.289~298 (2008)
- ML Wong, "Real-time PCR for mRNA quantitaion", BioTechniques, 39(1), pp.1~11 (2005)
- E Malinen, "Comparison of real-time PCR with SYBR Green I or 5'-nuclease assays and dot-blot hybridization with rDNA-targeted oligonucleotide probes in quatification of selected faecal bacteria", Microbioloy, 149, pp.269~277 (2003)
- P Kroken, "Quantifying Neodiprion sertifer nucleopolyhedrovirus DNA from insects, foliage and forest litter using the quantitative real-time polymerase chain reaction", Agrc.For.Entomol, 15, pp.120~125 (2013)
- G Munchow, "Automated chip-based device for simple and fast nucleic acid amplification", Expert Rev Mo Diagn, 5, pp.613~620 (2005)
- KE Herold, "Fabrication and Microfluidics" (2009), Caister Academic Press
- DA Mair, "Injection molded microfluidic chips featuring integrated interconnects", Lab Chip, 6, pp.1346~1354 (2006)
- SH Lee, "Rapid Detection of Mycobacterium Tuberculosis Using a Novel Ultra-Fast Chip-Type Real-Time PCR System", CHEST, 146(5), pp.1319~1326 (2014)
- JP Conde, "Lab-on-a-chip systems for integrated bioanalyses", PORTLAND PRESS, 60, pp.121~131 (2016)
- Asing, Eaqub Ali, "Lab-on-a-Chip-Based PCRRFLP Assay for the Detection of Malayan Box Turtle (Cuora amboinensis) in the Food Chain and Traditional Chinese Medicines", PLOS ONE, 11(10), pp.1~27 (2016)
- SL Weinrich, "Reconstitution of human telomerase with the template RNA component hTR and the catalytic protein subunit hTRT", Nat Genet, 17(4), pp.498~502 (1997)
- KL Kirkpatrick, "The significance of human telomerase reverse transcriptase (hTERT) in cancer", Eur J Surg Oncol, 27(8), pp.754~760 (2001)
- A Razzak, "A lab-on-achip-based mulitplex platform to detect fraud of bringing pig, dog, cat, rat and monkey meat in food chain", Food Adiit Contam, 32(11), pp.1902~1913 (2015)