RESUMO
Molecular clock analyses are challenging for microbial phylogenies, due to a lack of fossil calibrations that can reliably provide absolute time constraints. An alternative source of temporal constraints for microbial groups is provided by the inheritance of proteins that are specific for the utilization of eukaryote-derived substrates, which have often been dispersed across the Tree of Life via horizontal gene transfer. In particular, animal, algal, and plant-derived substrates are often produced by groups with more precisely known divergence times, providing an older-bound on their availability within microbial environments. Therefore, these ages can serve as "standard candles" for dating microbial groups across the Tree of Life, expanding the reach of informative molecular clock investigations. Here, we formally develop the concept of substrate standard candles and describe how they can be propagated and applied using both microbial species trees and individual gene family phylogenies. We also provide detailed evaluations of several candidate standard candles and discuss their suitability in light of their often complex evolutionary and metabolic histories.