Dna Methylation As Forensic Tissue Marker

Background and Objective The identification of tissue origins can be of crucial importance in forensic casework. Method for tiny and aged tissues discrimination remains a challenge due to many cross reaction existing in immunodetection and unstable mRNA. DNA methylation is an epigenetic mark on the mammalian genome. There are numerous tissue-dependent and differentially methylated regions (T-DMRs) in the unique sequences distributed throughout the genome. Recently study had shown that the detection tissue specific DNA methylation is a suitable technique to identify neuronal tissue in raw meat products which is prohibited by BSE (Bovine Spongiform Encephalopathy) legislation. Cell/tissue-specific expressions of various genes had been explained by the binding of cell/tissue specific transcription factors to their promoters. Taken together, in order to support a model in which the promoter regions special methylate pattern of gene are accessible for discriminate one tissue from the other. To study the feasibility of DNA methylation as forensic tissue markers, the promoter of human GFAP and DDX4 gene were selected as samples.Method Methylation levels of GFAP and DDX4 promoter of human tissue samples including brain, heart, lungs, liver, pancreas, spleen, kidney, skin, and semen(n=6) were detected by the combined bisulfite restriction enzyme analysis (COBRA), a semiquantitative DNA methylation analytical method. One-way variance of data was carried out and discrimate values were defined.Result The results showed that the methylation level of GFAP promotor in all huamn brain were lower than 45.82%((X|￣)_brain=44.70%, S_brain=8.38%), while that in all non-brain human tissues were larger than 58.60%((X|￣)_non-brain=68.01%, S_non-brain=1.42%),; When the discriminant value was set at 52.87%, brain and non-brain huamn tissues can be discriminated effectively. The methylation level of DDX4 promotor in all semen were lower than 50.41 % ((X|￣)_sperm=11.52%_EcoRI, 11.88%_TaqI ; S_sperm=8.98%_EcoRI, 10.25%_TaqI), while that in all somatic tissues were larger than 75.41 % ((X|￣)_somatic=82.73%_EcoRI, 82.94%_TaqI ; S_somatic=3.54%_EcoRI, 3.91%_TaqI). When the discriminant value was set at 49.85%(EcoRIsite)or 50.58%(TaqI site), semen and non- semen huamn tissues can be discriminated effectively.Conclusion Methylation level of GFAP promoter in brain tissue(including gray and white matter) was significantly lower than that in non-brain tissues, DNA methylation level of GFAP promoter can be used as a brain tissue-specific markers.Methylation level of DDX4 promoter in semen was significantly lower than that in somatic tissue, DNA methylation level of DDX4 promoter can be used as a semen-specific markers.As a proof of concept study, the sample size, methylation marker selected and techniques used here may not be suitable for the forensic practice. However, considering the characteristics of forensic samples and the advantages of DNA markers, tissue specific methylation-variable positions (MVPs) may be ideal tissue makers. Screening for different methylation regions (DMR) among tissues, developing a universally adapted DNA methylation-based tissue ID system may be a solution to the present problems in forensic tissue identification.