The non-overlapping RefSeq exons were then extended 90 bps on both ends and the human-mouse alignments [ 21] of the extended exons were extracted from the chained and netted human-mouse alignments in assembly hg12.
The potential non-exons in human genome were obtained by eliminating the alignment segment of human mRNAs and ESTs and the 100 bps beyond their end points from the chained and netted human-mouse alignments in assembly hg12, where the coordinates of human mRNAs and ESTs were based on the annotations of mRNA and ESTs in assembly hg12.
As shown in Figure 4A, the average rate of non-synonymous changes (dN) observed in human mouse alignments is 51% higher in the most closed chromatin regions of the genome than in the most open regions.
To test this, we compared synonymous (dS) and nonsynonymous (dN) substitution rates in human-mouse alignments for codons overlapping and non-overlapping with CGIs.
Predicted CpG islands acquired from the UCSC Genome Browser are shown in green, and chained human-mouse alignments are shown in olive green.
The (A)n simple repeat we found downstream of human and chimp putative -115 κB sites is absent in dog, mouse and rat (alignments in the additional files 2 and 3).
We downloaded the human-mouse alignments (hg17 vs mm5, [ 51]) for the human conservation curve in Figure 3 and mouse-human alignments for the mouse curve in Figure 4 (mm5 vs hg17, [ 51]).
UCSC alignments, on the other hand, are based on blastz pairwise alignments, which are not symmetric – a human-mouse alignment is different from a mouse-human alignment in general.
On imposing these filters we obtained 804311, 56062307 and 904717 alignments in human, mouse and rat respectively.
Of these approximately 200 SNPs, 36 were predicted to influence TF binding, in regions of sequence conservation of over 70% in human-mouse; the alignments for two such regions are indicated in additional files [See Additional file 2].
Babak et al. [ 38] used the QRNA algorithm [ 39] to search for ncRNAs in human-mouse pairwise alignments from intergenic and intronic regions conserved between human and mouse and rat.
