Fawcett et al. BMC Genomics 2011, 12:311 http://www.biomedcentral.com/1471-2164/12/311 [17,18]. Clearly, more information about genetic varia- tion segregating within this otherwise widely studied species would be useful. Additional effort to identify both common and low frequency genetic variation in this species is warranted, based on the potential opportunity for and significance of developing new genetic models of human disease. Despite the limited information available today, a num- ber of studies have shown that rhesus monkeys exhibit intra-species genetic variation in the same genes that have been associated with risk of disease in humans. Furthermore, recent analyses have demonstrated that DNA sequence variation among rhesus macaques can parallel the functional effects of known variation in humans, providing remarkably similar and thus highly informative animal models of human genetic risk. In one example, researchers found natural mutations segre- gating among captive rhesus monkeys in two genes that had previously been associated with macular degenera- tion in humans [19]. The investigators found that, as in humans, the polymorphisms in macaques were asso- ciated with individual variation in the major risk factor for macular degeneration, demonstrating a parallel genetic effect in rhesus and establishing this as an important model of genetic predisposition to this dis- ease. Other examples of genetic variation among rhesus macaques that parallels human functional variation are known in the serotonin transporter [20,21] and the mu- opioid receptor [22]. Based on these observations, we anticipated that gen- ome-wide knowledge of SNP variation segregating within research colonies of Indian-origin rhesus maca- ques would produce novel and valuable information about functionally significant genetic variation in a num- ber of important cellular pathways. Broad knowledge of both common and low frequency sequence variation in rhesus macaques (and by implication, equivalent infor- mation for other species of nonhuman primates) will facilitate discovery of new animal models of functional variation in specific genetic pathways and thus help investigators to elucidate mechanisms associated with human disease [17]. It is important to recognize that nonhuman primates constitute a unique set of animal models that offer a singular combination of research advantages. The overall genetic and physiological simi- larity of macaques and other nonhuman primates to humans [3,23] coupled with the opportunity to examine sets of animals raised under controlled environmental circumstances, or subject to precise environmental manipulations (e.g. exposure to therapeutic drugs or changes in diet), creates outstanding research opportu- nities. Analyses in macaques can examine the specific effects of individual genetic variants on those cellular pathways, the effects of genotype-by-genotype or Page 2 of 13 genotype-by-environment interaction, or other complex processes that are relevant to human disease but cannot be readily examined in human subjects under such tightly controlled environments. Unfortunately, little is known about genetic variation in rhesus macaques. Fewer than 900 validated SNPs and 100 CNVs have been described. The available data [23-28] suggest that a given number of rhesus macaques will show as much or more diversity and heterozygosity as the same number of humans. We undertook this study to assess the value of whole genome re-sequen- cing in rhesus macaques for the discovery of novel SNPs. We identified and bioinformatically validated SNPs throughout the rhesus macaque genome using existing sequence data and new SOLiD sequencing reads we generated from three unrelated macaques. We are particularly interested in the identification of novel variation among these animals that has potentially sig- nificant effects on gene function and physiological pro- cesses relevant to human disease. Results We identified and validated more than 3 million unique SNPs in rhesus macaques through pairwise comparisons of SNP data sets (Table 1). Our starting point for these analyses was a list of 4.3 million inferred heterozygous positions in the original Sanger sequencing data for the reference animal [23]. Comparison of these Sanger data with corona_lite SNP calls from SOLiD re-sequencing of the reference animal resulted in validation of 1,056,266 heterozygous positions (Figure 1). Similarly, re-sequen- cing of two additional Indian-origin animals, followed by comparison to the reference animal SNP list, resulted in the validation of an additional 873,552 SNPs. We were able to validate 1,058,581 additional SNPs among just three rhesus macaques using e-genotyping (http://is04607.com/~drio/egenotype/[29]). E-genotyping is a novel in silico method to survey mapped read sequence data using 31bp probe sequences created from a list of known or suspected SNPs. Unvalidated potential heterozygous positions from the original Sanger refer- ence animal data were used to create uniquely mapping e-genotyping probes. Using these probes we screened each of the SOLiD read sets (i.e. the reference animal and the two additional animals). This identified 527,425 validated SNPs that were not validated using the Sanger data and corona_lite analyses of SOLiD data. Probes cre- ated from the corona_lite analyses of SOLiD data were used to identify and validate 531,156 more SNPs. Finally, we used the smaller published datasets to complete a comprehensive list of validated SNPs. We validated 49,747 SNPs by comparing the results from the corona_lite re-sequencing analyses with previously published datasets. A small number of SNPs (n = 20)