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The relatively high coverage (x51 - x86) allowed testing the reproducibility of the alignment and thereby excluding alignment errors and sequencing errors over real changes.
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After excluding alignments with an e-value <10-5, two two best aligning, but different transcript models were implemented into the annotation table.
Additionally, likelihood bootstrap values from the reduced dataset obtained by excluding alignment-ambiguous regions are indicated.
Additionally, maximum parsimony bootstrap values from the reduced dataset obtained by excluding alignment-ambiguous regions are indicated.
Excluding alignment-ambiguous regions, however, does not take into account that different possible alignment solutions do not necessarily imply different topologies [ 33] or support values.
If this view is correct, the benefits of interspecies comparisons of affinity density outweigh the obvious shortcomings from excluding alignments.
After excluding alignments with unexpected stop codons, lengths less than 150 bp or Ks values above 0.1, 6,654 pairs of orthologs were retained for subsequent analysis.
In order to determine whether excluding alignments with lower MQ values would influence the results, sequencing depth and SNP statistics were also calculated for 1,223,938 reads with a MQ ≥ 30.
The whole set of somatic variants was manually checked within the BAM file in order to exclude alignment errors that in most cases occur in repetitive regions.
Alignments were trimmed with trimAl (Capella-Gutiérrez et al. 2009) with the parameter "-gt 0.8" to exclude alignment columns with gaps presented in more than 20% of the sequence.
This policy allows so called 'multireads' from genes with multiple copies to be reported, but excludes alignments to low-complexity sequence, to which failed reads often align.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com