Multiple alignments for each family were obtained by first computing the pairwise global alignment with the reference sequence using the gap program [ 55], and second by stacking all pairwise alignments to obtain a multiple alignment.
Multiple alignments for each family were obtained by first computing the pairwise global alignment with the reference sequence using the gap program [ 51], and then stacking all pairwise alignments to obtain a master-slave multiple alignment.
BLASTX programs, although capable of insightful results thanks to the six frame translations, have the limitation of not being able to transparently manage frameshifts that occur inside the sequence, for example by reconstructing an alignment from pieces obtained on different reading frames.
Note that a duplication event involves a single local alignment because alignments split by insertion of repeats are chained right after obtaining LASTZ alignments by a preprocessing step of the CAGE pipeline (Song et al. 2010).
Percentage identities between homologous genes were obtained by alignment of the proteins from whole genomes using the tBLASTn program [ 64].
The replacement values for the gap-free alignment regions were not changed significantly from the result obtained by alignment of full sequences including gaps.
The partition of genomic fragments in exons and introns was obtained by alignment of the genomic fragments with publicly available mRNA sequences.
Subsequently, SCGB1A1 and SCGBA1A cDNA consensus sequences were obtained by alignment of the 20 gene-specific sequences (accession numbers JQ906259, JQ906260, JQ906261, Additional file 1: Figure S1).
Using ∼12,000 particles, we obtained a 3D map by refining the alignment of the two AAA+ rings only, but then including the whole molecule in the reconstruction, which was at ∼17 Å resolution.
A similar number of putative novel gene loci (4431) was obtained by a reference alignment and prediction approach starting from the same dataset [ 33] (data not shown).
