Computational protocol: Testing the Role of Genetic Background in Parallel Evolution Using the Comparative Experimental Evolution of Antibiotic Resistance

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Protocol publication

[…] To obtain an accurate phylogeny of the genus Pseudomonas, we retrieved the nucleotide sequences of 55 housekeeping genes (supplementary table S1, Supplementary Material online) for 28 Pseudomonas genomes (supplementary fig. S1, Supplementary Material online) plus Azotobacter vinelandii from the Integrated Microbial Genomes database () using reciprocal BLAST to make sure of their orthology. Multiple alignments for all genes were obtained using MUSCLE () and afterwards trimmed with GBlocks () and concatenated with SequenceMatrix (). The resulting supermatrix was then used to infer the nucleotide substitution model using JModelTest 2 (). A maximum-likelihood (ML) phylogenetic tree was then inferred assuming a GTR+G+I model with PHYML (). Additional trees inferred using Bayesian inference and Neighbor Joining as well as a ML tree obtained with a supermatrix of the protein sequences yielded identical topologies (data not shown). The relevant phylogeny for the eight strains in this study system was then manually extracted. Branch lengths are proportional to genetic distance and all nodes on this phylogeny are strongly supported with at least 99% confidence, as determined by an approximate log-likelihood ratio test (Furio V, Maclean RC, unpublished manuscript). […]

Pipeline specifications

Software tools MUSCLE, Gblocks, Sequence Matrix, jModelTest, PhyML
Applications Phylogenetics, Nucleotide sequence alignment
Chemicals Rifampin