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Taxon and character sampling are central to phylogenetic experimental design; yet, we lack general rules. Goldman introduced a method to construct efficient sampling designs in phylogenetics, based on the calculation of expected Fisher information given a probabilistic model of sequence evolution. The considerable potential of this approach remains largely unexplored. In an earlier study, we applied Goldman's method to a problem in the phylogenetics of caecilian amphibians and made an a priori evaluation and testable predictions of which taxon additions would increase information about a particular weakly supported branch of the caecilian phylogeny by the greatest amount. We have now gathered mitogenomic and rag1 sequences (some newly determined for this study) from additional caecilian species and studied how information (both expected and observed) and bootstrap support vary as each new taxon is individually added to our previous data set. This provides the first empirical test of specific predictions made using Goldman's method for phylogenetic experimental design. Our results empirically validate the top 3 (more intuitive) taxon addition predictions made in our previous study, but only information results validate unambiguously the 4th (less intuitive) prediction. This highlights a complex relationship between information and support, reflecting that each measures different things: Information is related to the ability to estimate branch length accurately and support to the ability to estimate the tree topology accurately. Thus, an increase in information may be correlated with but does not necessitate an increase in support. Our results also provide the first empirical validation of the widely held intuition that additional taxa that join the tree proximal to poorly supported internal branches are more informative and enhance support more than additional taxa that join the tree more distally. Our work supports the view that adding more data for a single (well chosen) taxon may increase phylogenetic resolution and support in weakly supported parts of the tree without adding more characters/genes. Altogether our results corroborate that, although still underexplored, Goldman's method offers a powerful tool for experimental design in molecular phylogenetic studies. However, there are still several drawbacks to overcome, and further assessment of the method is needed in order to make it better understood, more accessible, and able to assess the addition of multiple taxa.

In molecular phylogenetic studies, a major aspect of experimental design concerns the choice of markers and taxa. Although previous studies have investigated the phylogenetic performance of different genes and the effectiveness of increasing taxon sampling, their conclusions are partly contradictory, probably because they are highly context specific and dependent on the group of organisms used in each study. Goldman introduced a method for experimental design in phylogenetics based on the expected information to be gained that has barely been used in practice. Here we use this method to explore the phylogenetic utility of mitochondrial (mt) genes, mt genomes, and nuclear rag1 for studies of the systematics of caecilian amphibians, as well as the effect of taxon addition on the stabilization of a controversial branch of the tree. Overall phylogenetic information estimates per gene, specific estimates per branch of the tree, estimates for combined (mitogenomic) data sets, and estimates as a hypothetical new taxon is added to different parts of the caecilian tree are calculated and compared. In general, the most informative data sets are those for mt transfer and ribosomal RNA genes. Our results also show at which positions in the caecilian tree the addition of taxa have the greatest potential to increase phylogenetic information with respect to the controversial relationships of Scolecomorphus, Boulengerula, and all other teresomatan caecilians. These positions are, as intuitively expected, mostly (but not all) adjacent to the controversial branch. Generating whole mitogenomic and rag1 data for additional taxa joining the Scolecomorphus branch may be a more efficient strategy than sequencing a similar amount of additional nucleotides spread across the current caecilian taxon sampling. The methodology employed in this study allows an a priori evaluation and testable predictions of the appropriateness of particular experimental designs to solve specific questions at different levels of the caecilian phylogeny.