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Published on: June 29, 2020
Novel identification of biofluids using a multiplex methylation sensitive restriction enzyme-PCR system
Yu-Chih Lin1, Li-Chin Tsai2, James Chun-I Lee3
1Graduate Institute of Biotechnology, National Chung Hsing University, 250 Kuo Kuang Road, South District, Taichung City 40227, Taiwan, ROC; Taichung City Government Police Department, 588 Wenxin Road Section 2, Xitun District, Taichung City 40758, Taiwan, ROC.
Forensic science benefits from a new 10-plex methylation sensitive restriction enzyme-PCR (MSRE-PCR) system. This method accurately identifies body fluids like blood and saliva from DNA samples in a single reaction.
Area of Science:
- Forensic science
- Molecular biology
- Genetics
Background:
- Accurate identification of body fluids is critical in forensic investigations.
- DNA methylation profiles offer a promising avenue for biofluid discrimination.
- Existing methods may lack specificity or require multiple analyses.
Purpose of the Study:
- To develop and validate a multiplex methylation sensitive restriction enzyme-PCR (MSRE-PCR) system for body fluid identification.
- To identify novel DNA methylation markers specific to various human biofluids.
- To integrate biofluid identification with human DNA profiling in a single assay.
Main Methods:
- Searched the Infinium HumanMethylation450 BeadChip database for biofluid-specific methylation markers.
- Developed a 10-plex MSRE-PCR assay combining 8 biofluid-specific markers and 2 control markers.
- Analyzed 100 DNA samples from five body fluid types (semen, vaginal fluids, saliva, venous blood, menstrual blood).
- Validated the assay for sensitivity, reproducibility, and performance with mixed body fluids.
- Performed co-amplification with AmpFlSTR® MiniFiler™ PCR Amplification Kit for simultaneous DNA profiling.
Main Results:
- The 10-plex MSRE-PCR system generated unambiguous results for identifying the origin of DNA samples from the five tested body fluid types.
- Validation studies confirmed the assay's sensitivity, reproducibility, and ability to handle mixed body fluid samples.
- Simultaneous generation of methylation profiles and miniSTR profiles was achieved in a single reaction.
Conclusions:
- The developed 10-plex MSRE-PCR system is a powerful and applicable tool for body fluid identification in forensic science.
- This integrated approach allows for simultaneous human identification and body fluid typing, enhancing forensic casework efficiency.
- The findings support the utility of DNA methylation analysis for forensic biofluid discrimination.
