Development of a Model for Predicting Chronological Age Based on DNA Methylation Data from Buccal Swabs Using Ion Torrent S5 Technology

Age estimation using DNA methylation analysis is one of the tools currently used in forensic science to facilitate human identification. A total of 117 buccal swab samples from individuals aged 18 to 67 years were analyzed using a custom panel encompassing 610 CpG sites across 12 age-related genes. The analysis was performed using the amplicon-based technology Ion AmpliSeq™, which incorporates Ion Torrent S5 sequencing (Thermo Fisher Scientific, Waltham, MA, USA). The resulting methylation data were used to develop and compare age-prediction models based on Watson-strand, Crick-strand, and combined Watson + Crick methylation matrices. After candidate CpG selection and model reduction, the final Watson-strand model used 5 CpG sites from the age-related genes C1orf132, ELOVL2, KLF14, PDE4C and TRIM59 and was fitted using Elastic Net regression with alpha grid search, lambda selection and internal cross-validation. Model validation yielded prediction results with a mean absolute error of 4.42 years for the Watson matrix, 4.75 years for the Crick matrix, and 5.08 years for the WC matrix. These results allow for the evaluation of the suitability of using Ion AmpliSeq™ technology in forensic age estimation studies based on DNA methylation analysis.