Diamond™ Nucleic Acid Dye를 사용한 Touch DNA 시각화 및 검출
Visualization and Detection of Touch DNA using Diamond™ Nucleic Acid Dye
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초록
본 연구에서는 범죄현장 및 증거물에 잠복되어 있는 DNA (Deoxyribose Nucleic Acid)를 시각화하여 정확한 위치에서 유전자 증거물을 채취하고자 젤 전기영동에 사용되는 핵산 염색 시약 중 하나인 Diamond™ Nucleic Acid Dye를 사용하였다. 사람이 손으로 만진 물품의 표면에 Diamond™ Nucleic Acid Dye (10,000X) 원액을 75% 에탄올로 20배 희석하여 염색하였다. 생물학적으로 근연관계가 없는 4명의 공여자에게 멸균 처리된 물품을 만지게 한 후, 20배 희석한 DD(Diamond™ Nucleic Acid Dye)를 물품의 표면에 피펫으로 점적하여 Touch DNA를 염색하였다. 그리고 Dino-Lite 디지털 형광현미경을 사용하여 물품 표면의 형광을 확인하였다. DD 분자는 구조적으로 DNA의 골격 (sugar-phosphate backbone)에 효과적으로 결합할 수 있기 때문에 ssDNA (single-stranded DNA), dsDNA (double-stranded DNA) 그리고 RNA를 형광으로 확인 할 수 있다. DD는 dsDNA를 기준으로 청색 파장(494nm)에서 최대 여기(excitation)를 가지며 DNA와 결합하면 녹색 형광(558nm)을 방출(emission)한다. 이로써 4명의 공여자가 만진 물품에서 녹색 형광을 방출하는 발광체를 관찰할 수 있었으며, 멸균 면봉에 멸균수를 묻혀서 녹색 형광 을 발현하는 발광체를 채취하였다. 현재 경찰청에서 국립과학수사연구원으로 유전자 감정 의뢰과정과 동일하게 사용 중인 유전자 증거물 채취 키트에 포장하여 한국유전자정보연구원으로 가져간 후 STR (Short Tandem Repeat) profiling 분석을 하였으며, 공여자 4명의 구강상피세포 DNA에서 분석된 STR profile을 대조한 결과 개인식별이 가능한 것을 확인하였다. 이 방법이 보급 된다면 보이지 않는 DNA를 시각화하여 유전자증거물을 채취할 수 있도록 과학수사관을 안내하고 DNA가 없는 시료의 무분별한 감정 처리 시간 단축 및 예산 절약의 효과가 있다고 사료되었다.
Abstract
This study used Diamond™ Nucleic Acid Dye, a nucleic acid staining solution used for gel electrophoresis to precisely locate and collect genetic evidence from the crime scene and evidence by visualizing DNA (Deoxyribose Nucleic Acid). Diamond™ Nucleic Acid Dye (10,000X) was diluted by 20times with 75% ethanol and was used to dye the surface of the touched items. After having four biologically unrelated donors touch sterilized items, the diluted Diamond™ Nucleic Acid Dye(DD) solution was dripped onto the surface of the items using a pipette to stain the Touch DNA. A dino-Lite digital fluorescence microscope was then used to observe fluorescence. ssDNA(single-stranded DNA), dsDNA(double-stranded DNA) and RNA can be confirmed with fluorescence since the structure of DD molecules effectively combine DD molecules with the sugar-phosphate backbone of the DNA. With dsDNA at the baseline, DD has the greatest excitation at the blue wavelength(494nm) and emits green fluorescence(558nm) when combined with DNA. Green fluorescence was observed on the items touched by the four donors, and sterilized cotton swabs moistened with sterile water were used to collect luminous matter emitting green fluorescence. To replicate the current genetic analysis request procedure, the sample was placed in the genetic evidence collection kit, sent to the Korea genome Information Institute from the police agency, and analyzed with STR(Short Tandem Repeat) profiling. A comparison with the STR profiles derived from the buccal epithelial cells of the four donors confirmed that the collected sample could successfully identify the donors. Widespread use of the technique presented in this research is expected to visualize invisible DNA and guide forensic field agents, reduce the processing time of DNA-less staining solution and result in a budget-saving.
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