Specifically, we compare node dating using nine calibration points derived from the fossil record with total-evidence dating based on 343 morphological characters scored for 45 fossil (4--20 complete) and 68 extant taxa.
In both cases we use molecular data from seven markers (∼5 kb) for the extant taxa.
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Once the bones are done growing in length, that avenue is closed. Chemical analysis of tooth dentin, such as aspartic acid racemization, has shown reproducible and fairly precise results.
Spalding and her postdoc advisor Jonas Frisén had a hunch that a pulse of radioactive carbon created by above-ground nuclear tests during the Cold War could help solve the riddle.
“A geopolitical phenomenon—this Cold War bomb testing—has, in a way, put a date stamp on everything and everybody,” Spalding says.
These results emphasize the importance of considering markers from multiple genomes and alternative fossil placements when addressing evolutionary issues that depend on ages estimated for important groups. To allow additional calibration points and the comparison of evolutionary rates among all angiosperms, taxa outside the grasses were added to this initial data set as follows: the three selected coding genes were first retrieved from complete plastid genomes available in NCBI database; then additional taxa were added that had available sequence data for all three plastid regions such that the complete data set contained representatives for most angiosperm orders and most monocot families.
In the absence of an exceptionally good fossil record, divergence times must be inferred from genetic markers. Often, however, there is a low number of fossil calibration points relative to a large number of species (and thus nodes in the phylogeny). The whole data set was aligned with MUSCLE v3.6 (Edgar 2004) and the alignment was manually refined.