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To this end, we first estimated the ITS nrDNA substitution rate in BEAST (as described above) using the inferred dates when the two Nicotiana species became isolated on different Pacific Ocean islands.
Congruence between independent methods can greatly increase the confidence in the inferred dates.
The inclusion of additional more closely related outgroups could improve the accuracy of the inferred dates [ 52].
Regardless, violation of an implicit autocorrelation assumption did not have significant effects on inferred dates for nodes of interest (Table 3).
A macrofossil of an extant species Acacia melanoxylon identified from the Pliocene [ 27] enabled us to compare our inferred dates with that of the fossil record.
According to the inferred dates resulted from BEAST, the two main mitochondrial lineages of A. andreanskyi (South versus central and North) split approximately 7.6 Ma (95% high posterior density (HPD) interval 4.3-11.9 Ma).
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Additionally, dates of individual mutation events (epitope, non-epitope, and synonymous, each with a distinct symbol) could be marked on the right hand side based on their inferred date, so that readers can see the raw data.
Thus, we made the decision to restrict ourselves to assuming a single-point admixture, fit for a principled threshold d > d0, accepting that the inferred date n represents some form of average value over the true history.
As an independent check of our inferred date estimates, we estimated rates of mitochondrial evolution within Crenadactylus using posterior age estimates from the nuclear and two different combined analyses.
The inclusion of fossils as tip taxa thus clearly affects inferred divergence dates, and suggests that the early divergence dates for the most species-rich modern clade of snakes are younger than previously assumed.
These inferred hatching dates are a month earlier than the hatching dates recorded at Ice Lake in 1993 [ 35].
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