Genome-wide methylation profiling and a multiplex construction for the identification of body fluids using epigenetic markers.

The identification of body fluids found at crime scenes can contribute to solving crimes by providing important insights into crime scene reconstruction. In the present study, body fluid-specific epigenetic marker candidates were identified from genome-wide DNA methylation profiling of 42 body fluid samples including blood, saliva, semen, vaginal fluid and menstrual blood using the Illumina Infinium HumanMethylation450 BeadChip array. A total of 64 CpG sites were selected as body fluid-specific marker candidates by having more than 20% discrepancy in DNA methylation status between a certain type of body fluid and other types of body fluids and to have methylation or unmethylation pattern only in a particular type of body fluid. From further locus-specific methylation analysis in additional samples, 1 to 3 CpG sites were selected for each body fluid. Then, a multiplex methylation SNaPshot reaction was constructed to analyze methylation status of 8 body fluid-specific CpG sites. The developed multiplex reaction positively identifies blood, saliva, semen and the body fluid which originates from female reproductive organ in one reaction, and produces successful DNA methylation profiles in aged or mixed samples. Although it remains to be investigated whether this approach is more sensitive, more practical than RNA- or peptide-based assays and whether it can be successfully applied to forensic casework, the results of the present study will be useful for the forensic investigators dealing with body fluid samples.