Epigenetic discrimination of identical twins from blood under the forensic scenario

Abstract

Monozygotic (MZ) twins share the same STR profile, demonstrating a practical problem in forensic casework. DNA methylation has provided a suitable resource for MZ twin differentiation; however, studies addressing the forensic feasibility are lacking. Here, we investigated epigenetic MZ twin differentiation from blood under the forensic scenario comprising i) the discovery of candidate markers in reference-type blood DNA via genome-wide analysis, ii) the technical validation of candidate markers in reference-type blood DNA using a suitable targeted method, and iii) the analysis of the validated markers in trace-type DNA. Genome-wide methylation analysis in blood DNA from 10 MZ twin pairs resulted in 19-111 twin-differentially methylated sites (tDMSs) per pair with >0.3 twin-to-twin differences. Considering all top three candidate tDMSs across all pairs in the technical validation based on methylation-specific qPCR, 67.85% generated >0.1 twin-to-twin differences. Of the validated tDMSs, 68.4% showed >0.1 twin-to-twin differences with qPCR in trace-type DNA across 8 pairs. Using an updated marker selection strategy, 8 additional candidate tDMSs were obtained for an example MZ pair, of which 7 showed >0.1 twin-to-twin differences in both reference- and trace-type DNA. Lastly, we introduce a high-resolution melting curve analysis of the entire fragment that can complement the proposed approach. Overall, our study demonstrates the general feasibility of epigenetic twin differentiation in the forensic context and highlights that the number of informative tDMSs in the final trace DNA analysis is crucial, as some candidate markers identified in reference DNA were shown not informative in the trace DNA due to various, including technical, reasons. Future studies will need to address the optimal number of epigenetic markers required for reliable identification of MZ twin individuals including statistical considerations.

From forensic epigenetics to forensic epigenomics: broadening DNA investigative intelligence

“…Although some studies have explored the value of epigenetic profiling in forensically discriminating MZ twins [ 62 , 63 ], it is not yet fully established whether the observed twin-to-twin differences are twin pair-specific, or might be universal and applicable across twin pairs, as would be preferred. Recently, a first attempt was made to demonstrate the feasibility of differentiating between MZ twins using forensic epigenetics [ 15 ]. This study showed that most, but not all, twin-differentiating CpG sites (which were identified using genome-wide screening technologies in reference-type blood DNA) could be replicated by targeted methods that are suitable for forensics in trace-type DNA from bloodstains, highlighting technical challenges [ 15 ].…”

“…Currently, only a limited number of DNA methylation markers are applied for a few forensic purposes, using technologies that enable the analysis of a small number of such markers. These approaches can be classified as forensic epigenetics, and include DNA methylation profiling for tissue determination [ 13 ], age prediction [ 14 ], and differentiation between monozygotic twins [ 15 ]. The concept of personalized epigenomics, which is already used in medical research [ 16 ], has not yet been recognized in the forensic field.…”

Identification of the perpetrator among identical twins using next-generation sequencing technology: A case report

“…However, it is not always effective in analyzing samples in regular criminal or paternity testing cases. Small genetic or epigenetic differences have been identified for distinguishing between MZ twins [2][3][4][5]. Additionally, extremely rare mutations in the nuclear genome, identified by whole-genome sequencing (WGS), were useful in distinguishing between identical twins in paternity testing [6].…”

Could Rapidly Mutating Y-STRS Be a Potential Forensic Tool in Discriminating Lebanese Monozygotic Twins?

“…Accurate DNA typing results are nowadays the basis of increasing number of judicial decisions (Vidaki et al, 2017). Still, the main limitation in forensic genetics involves distinguishing individuals within a pair of monozygotic twins in criminal or paternity cases (Stewart et al, 2015).…”

“…This scenario poses an urgent challenge to forensic genetics especially that monozygotic twinning accounts for 1 in 250 live births on average (Fraga et al, 2005). Till the present day, and despite the high power of discrimination that characterizes the well identified 16 to 24 autosomal short tandem repeat (STR) markers, monozygotic twins discrimination is still limited in forensics (Xu et al, 2015;Vidaki et al, 2017). Furthermore, lineage markers are in general less informative than autosomal ones.…”