Appendix 4.3.4: PAML-BEB selected site analysis

To validate our findings from aBSREL and MEME using a different method, we applied Phylogenetic Analysis by Maximum Likelihood (PAML) branch by site analysis to the luciferase sequences. We tested the alternative hypothesis, that there is a class of sites under selection (ω > 1) on the EAncLuc ancestral branch identified as under selection in the aBSREL analysis, against the null hypotheses, that all classes of sites on all branches are evolving either under constraint (ω < 1) or neutrality (ω = 1). A likelihood ratio test supported the alternative hypothesis, that 13% of sites in luciferase were in a positively selected class (ω = 3.25). Subsequent Bayes Empirical Bayes (BEB) estimation identified 31 sites with evidence of selection on these branches, 5 of which were significant.