Evaluating Phylogenetic Inference with AliGROOVE

To test the efficiency of AliGROOVE we designed two sets of nucleotide and amino acid sequence data using 4-taxon and 6-taxon trees (Figure 1). The topology of the 4-taxon setup (setup A, Figure 1a) contained two long branches of unrelated taxa (with branch lengths BL 2 = 0.1,0.3,0.5,0.7,0.9,1.1,1.3,1.5) under three different branch length conditions for the other two short terminal branches (BL 3 = 0.1,0.12,0.14 and RB = 0.1) and two different lengths of the short internal branch (BL 1 = 0.01,0.02). The 6-taxon setup (setup B, Figure 1b) contained two long internal branches (BL 2 = 0.1,0.3,0.5,0.7,0.9,1.1,1.3,1.5), separated by a short internal branch (BL 1 = 0.01) while the lengths of terminal branches are kept constant (BL 3 = 0.01 and RB = 0.1). For both test setups, 100 alignments were generated for each step of BL 2 branch elongation. Sequence length of each alignment of setup A was set to 250,000 character state positions and for setup B to 50,000 character state positions to reduce the calculation time. All alignments were generated with INDELible v.1.03 [24 (link)]. In order to simulate nucleotide sequence data we used the Jukes-Cantor model (JC) of sequence evolution and for amino acid sequence data the BLOSUM62 substitution model. All data were simulated with among site rate variation (ASRV), using a mixed-distribution model with a shape parameter α = 1.0, and a proportion of invariant sites ρ_inv= 0.3. ASRV was modelled using a continuous Γ-rate distribution while indel events were not simulated.
Trees of simulated data were inferred with PhyML_3.0_linux64 [25 (link), 26 (link)]. We analyzed the data with a mixed-distribution model (JC+ Γ + I) and correct parameter values (α = 1.0, ρ_inv= 0.3), except for the categorization of the gamma distribution. The number of relative substitution rate categories was set to four (c = 4) and tree topologies and branch lengths were optimized. Maximum Likelihood analyses were performed and evaluated with a Perl pipeline. For each branch length-combination, we generated 100 data replicates and recorded the frequencies of correct and incorrect tree reconstructions using correct alignments and nearly correct substitution models (Figures 2, 3, Additional files 1, 2, 3).