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To reduce the number of false indels arising from wrong alignments, we employed sequence alignment programs based on structures to perform pairwise alignments of non-redundant structures in each family.
Thus, we reconstructed the sequences of the human chimp ancestor promoters employing multiple sequence alignments of human, chimp, and rhesus (see Material and Methods).
Subsequently, MEGA version 6.06 was also employed for multiple sequence alignment of the amino acids.
The widely used SIFT algorithm (Sorting Intolerant From Tolerant) employs a multiple sequence alignment of homologous proteins to identify conserved regions in the protein of interest, each possible substitution can then be scored according to the conservation observed at each position (Ng and Henikoff 2001).
In order to analyze ST repeats, WebLogo (http://weblogo.berkeley.edu/) and Two Sample Logo (TSL) (http://www.twosamplelogo.org/) were employed as graphical representation for multiple sequence alignment of conserved positions of the amino acid in the repeats [ 47].
PhastCons score employed in this study provides an estimate of the individual nucleotide level conservation, calculated based on multiple sequence alignment of 46 vertebrate genomes with respect to mouse reference genome mm10.
A multiple sequence alignment of the amino acid sequences of all available arthropod hemocyanins and selected arthropod phenoloxidases was constructed employing MAFFT 6 [ 33] with the G-INS-i routine and the BLOSUM 45 matrix at http://mafft.cbrc.jp/alignment/server/.
Sequence alignment of Piezo orthologues.
(b) Multiple sequence alignment of NucS sequences.
b, Sequence alignment of TcB sequences.
c, Sequence alignment of TcB sequences.
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