SupplementalMethods.docVIP

Supplemental Methods Translated cDNA sequences and amphioxus genome release v1.0 gene models were aligned to published deuterostome, cnidarian (Nematostella vectensis), and Drosophila melanogaster protein sequences using ClustalX [1]. Bootstrapped Neighbor-Joining trees [2] were then constructed using ClustalX, and drawn using NJplot [3]. Bootstrap values were calculated from 1000 resamplings of the alignment data. Maximum Likelihood trees and quartet-puzzling reliability scores were generated from the same alignments using TREE-PUZZLE [4]. For most trees, cnidarian or Drosophila sequences served as outgroups. For others, related non-homologous deuterostome proteins were chosen. Supplemental Figure Legends Figure S1. Phylogenetic analyses of amphioxus Alx and Bapx. (A) Bootstrapped Neighbor-Joining tree of deuterostome and Drosophila Alx homologs. The related Drosophila retinal homeobox (Rx) gene serves as an outgroup. (B) Maximum Likelihood tree generated using the same alignment as in (A). (C) Bootstrapped Neighbor-Joining tree of vertebrate, amphioxus, Drosophila and cnidarian Bapx/Bagpipe homologs. The cnidarian Bapx homolog NK3 serves as the outgroup. (D) Maximum Likelihood tree generated using the same alignment as in (C). In all analyses the amphioxus gene groups strongly with its vertebrate or invertebrate deuterostome homologs, supporting orthology. Bootstrap values and quartet-puzzling reliability scores are shown at the branch points, and distance units are shown in the upper right hand or lower left hand corners of the trees. Gene names are prefixed by the initials of their respective species names. Species included in the analyses are Drosophila melanogaster, Nematostella vectensis, Strongylocentrotus purpuratus, Paracentrotus lividus, Branchiostoma floridae, Danio rerio, and Mus musculus. Figure S2. Phylogenetic analyses of amphioxus Barx, BarH, and Ets. (A) Bootstrapped Neighbor-Joining tree of deuterostome and Drosophila Barx and Ba