Abstract

Summary: Ancient DNA studies are increasingly targeting samples that are beyond the limit of radiocarbon dating (>50 thousand years old) and are often difficult or impossible to date using other geochronological methods. In cases where complete mitochondrial genomes (mitogenomes) can be recovered from such samples, Bayesian molecular clock dating approaches are routinely used as an alternative method for estimating their age. However, molecular clock dating of ancient mitogenomes lacks a standardised, reproducible computational framework, and existing approaches rely heavily on graphical interfaces that limit automation and scalability. To address these gaps, we developed MitoDate, an automated Nextflow pipeline for reproducible molecular clock dating of ancient mitochondrial genomes. The workflow standardises Bayesian time-calibrated phylogenetic inference within a portable, containerised framework, reducing manual intervention and improving analytical consistency. Availability and implementation: MitoDate is implemented in Nextflow and is freely available at https://github.com/CpgSthlm/MitoDate. The pipeline is distributed with containerised dependencies and detailed documentation, including example datasets and usage guidelines.