ena-STUDY-CNRS UMR 5288-18-03-2021-09:07:50:737-1860
Performance and automation of ancient DNA capture with RNA hyRAD probes.
DNA hybridization-capture techniques allow to focus sequencing efforts on pre-selected genomic regions. This feature is especially useful for characterizing ancient DNA (aDNA), since aDNA extracts are often dominated by environmental sources instead of the focal species. However, most of the experimental capture procedures available to aDNA research rely on commercially synthesized probes, which can prove expensive and are designed from the sequence variation present in well-characterized genome panels. Here, we explored for the first time the efficacy of a modified hyRAD protocol on osseous aDNA as an inexpensive and design-free alternative to commercial capture protocols. While enrichment was limited for those samples already characterized by good DNA preservation, aDNA extracts with low endogenous content (<1%) could be enriched up to 53-fold. The aDNA sequences mapped preferentially on the genomic regions targeted by the probes, showing on-target enrichment of up to 146-fold for those samples limited in endogenous DNA. Performing two rounds of capture instead of a single round increased the capture specificity and increased on-target coverage up to 3.60-fold. The versatility of the hyRAD approach, in particular the possibility to use methylation sensitive restriction enzymes, allowed us to produce probes targeting hypomethylated regions that  are associated with lower postmortem DNA damage rates and  can, thus, deliver higher sequence quality. Finally, we developed a fully automated hyRAD protocol using inexpensive robotic platform. Overall, our work established hyRAD as a cost-effective strategy in ancient population genomics to recover a set of shared orthologous variants across multiple samples.
DNA hybridization-capture techniques allow to focus sequencing efforts on pre-selected genomic regions. This feature is especially useful for characterizing ancient DNA (aDNA), since aDNA extracts are often dominated by environmental sources instead of the focal species. However, most of the experimental capture procedures available to aDNA research rely on commercially synthesized probes, which can prove expensive and are designed from the sequence variation present in well-characterized genome panels. Here, we explored for the first time the efficacy of a modified hyRAD protocol on osseous aDNA as an inexpensive and design-free alternative to commercial capture protocols. While enrichment was limited for those samples already characterized by good DNA preservation, aDNA extracts with low endogenous content (<1%) could be enriched up to 53-fold. The aDNA sequences mapped preferentially on the genomic regions targeted by the probes, showing on-target enrichment of up to 146-fold for those samples limited in endogenous DNA. Performing two rounds of capture instead of a single round increased the capture specificity and increased on-target coverage up to 3.60-fold. The versatility of the hyRAD approach, in particular the possibility to use methylation sensitive restriction enzymes, allowed us to produce probes targeting hypomethylated regions that  are associated with lower postmortem DNA damage rates and  can, thus, deliver higher sequence quality. Finally, we developed a fully automated hyRAD protocol using inexpensive robotic platform. Overall, our work established hyRAD as a cost-effective strategy in ancient population genomics to recover a set of shared orthologous variants across multiple samples.