Whether or not DNA variation changes genome-wide nucleotide compositions remains largely unknown. By examining 4,604,291 DNA variants between two rat strains, we observed that sequencing depth is strongly correlated with genome content as 21.41, 38.36, 44.26 and 6512.70 average reads per locus were collected for Y, X, autosomes and mitochondrial (MT) genomes; respectively (P<0.0001). The mutation rates corresponding to these four genome subsets were 0.055, 0.401, 1.733 and 4.475 variants per kb (P<0.0001), confirming the links between recombination frequencies and DNA variability. Although SNPs (single nucleotide polymorphisms) tend to reduce AT content, more CG deletions than CG insertions (INDELs) implies the GC content would not increase. Therefore, the SNP-INDEL interplay may play a key role in maintenance of the AT-rich genomes in rat during evolution. Formation of CpG sites appear to be hindered because genome-wide G INDELs (1.38%) with C as the 5’-nucleotide and CG INDELs (1.19%) are rare. However, the relatively high C—>G/G—>C rate in 5’UTRs (untranslated regions) and G/C INDELs in the 5’UTR and/or exonic regions highlight their importance for execution of gene function. Our study provides evidence that DNA variation does not jeopardize genome stability and functional conservation during evolution.