DNA methylation analysis of ELOVL2 gene using droplet digital PCR for age estimation purposes

DNA methylation (DNAm) evaluation has been investigated in various tissues and body fluids allowing the identification of many CpG markers with high correlations with chronological age, potentially useful for forensic analysis. We developed a duplex droplet digital PCR (ddPCR) assay to address methylation levels of ELOVL2 gene CpG sites. DNA samples obtained from peripheral blood of 56 healthy individuals (35 women, 21 men; aged 1-94 years old) were submitted to bisulfite conversion, followed by ddPCR analysis for a selected region of ELOVL2 and the reference gene C-LESSC1. Simple linear regression revealed a strong correlation between DNAm simultaneous captured from five CpG sites of ELOVL2 gene and chronological age (R = 0.892; P = 2.84 × 10), explaining 79.2% of age variation. The obtained mean absolute deviation (MAD) between predicted and chronological ages was 10.07 years. Here we describe a ddPCR-based assay to assess DNAm in ELOVL2 gene as a biomarker for potential use in forensic age prediction.