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The multiple alignments were generated by PSI-Coffee, an aligner within the T-Coffee multiple alignment package that aligns distantly related protein sequences using homology extension [ 83, 84 ].
Multiple alignments were generated by the MULTALIN program (Corpet 1988).
The coevolved mutation scores from multiple sequence alignments were generated by PSICOV algorithm [12].
Multiple alignments were generated by MegAlign from the LaserGene software package using the CUSTALW algorithm.
Alignments were generated by three iterations of PSI-BLAST [66] searches against UniProt using our standard protocol for the generation of profiles [67].
All possible ungapped alignments were generated by shifting this block along the template sequence (Fig. 2), with the constraint that the SSE of the target sequence always contained a constant number of amino acids.
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The final alignments
The alignment was ordered from the most highly varied sites to the most conserved sites, and a series of alignments was generated by successively shortening the OV-sorted alignment in steps of 250 sites.
Each column of these alignments was generated by randomly selecting a statistical model (from the set of 5242 models) and then emitting 20 random amino acid characters with probabilities derived from the chosen model.
It works as follows [ 9, 6]: for a fixed alignment of sequences, a collection of new alignments is generated by deleting some columns from the alignment at random and replacing them by other columns from that alignment, also picked at random.
Maximum-likelihood trees based on the codon alignment were generated by RAxML v.7.0.3 and Garli v.1.0 where generalized time reversible (GTR) was used as the substitution model (Stamatakis 2006; Zwickl 2006).
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