BMC Genomics 2009, 10:582 http://www.biomedcentral.com/1471-2164/10/582 frequency in germplasm samples that have not been suffi- ciently inbred to reach a state of genome-wide allele fixa- tion. The spatial positions of heterozygote and homozygote data clusters were confined to areas of high certainty so that data points with less certainty fell outside the boundaries of heterozygotes and homozygotes and were scored as "no-call" (Fig 2A, one germplasm sample as black dot). Polymorphisms with theta compressed clus- ters were not used if the compression was such that any homozygote call was not plainly distinguishable (Figure 2B, set as Gentrain 0.000, 100% "no call"). Vertically sep- arated data clusters were not accepted as polymorphisms (Figure 2C). Following the production of one master workspace for each Pilot OPA using all DNA samples, cus- tomized workspaces were produced for each mapping population to optimize the genotype calls via minor adjustments of cluster positions. Genotype calls were exported as spreadsheets from BeadStudio and then parsed to create input for mapping programs. Individual and consensus map production Individual maps were made principally using MSTMap [29,30] for each data set from the four doubled haploid mapping populations. In brief, MSTMap first identifies linkage groups, then determines marker order by finding the minimum spanning tree of a graph for each linkage group, then calculates distances between marker using recombination frequencies. JoinMap 4 [31] was used to confirm linkage groups and marker order determined by MSTMap. Raw data for problematic markers were reviewed using BeadStudio and then either the marker was discarded entirely if any ambiguity in data calling could not be resolved or individual genotype calls were modified if it was plainly evident that such adjustments were warranted. Each such review of primary data was fol- lowed by the production of new maps; this iterative proc- ess generally involved 10-20 cycles for each individual map. At several points, a consensus map was produced using MergeMap [32], which also flags problematic mark- ers for review. MergeMap takes into account marker order from individual maps and calculates a consensus marker order. Briefly, the input to MergeMap is a set of directed acyclic graphs (DAGs) [33] from each individual map, and the output is a set of consensus DAGs (Figure 3, Fig- ures S3-S9, Additional Files 6, 7, 8, 9, 10, 11, 12), where each is consistent with all (or nearly all) of the markers in the individual input maps. MergeMap then linearizes each consensus DAG using a mean distance approximation. The consensus map coordinates from MergeMap were normalized to the arithmetic mean cM distance for each linkage group from the four individual maps (Figure S2, see Additional File 4 and Table S4, see Additional File 14). Implementation of BOPA1 and BOPA2 in US barley breeding germplasm As part of Barley CAP [17], the two BOPAs have been part of an effort to genotype a total of 3840 US barley breeding lines contributed from ten US barley breeding programs for association mapping analyses. As of January 2009, data from both BOPAs had been generated for 1920 breeding lines, with 960 submitted from the selections of each of two years, 2006 and 2007. Table S5 (Additional File 15) provides MAF for observed in these samples for each SNP in BOPA1 and BOPA2. Abbreviations AFLP: amplified fragment length polymorphism; DAG: directed acyclic graph; DArT: diversity array technology; EST: expressed sequence tag; QTL: quantitative trait locus; RFLP: restriction fragment length polymorphism; SNP: single nucleotide polymorphism; SSR: simple sequence repeat; STS: sequence tagged site. Authors' contributions The contributions of authors and other colleagues, loca- tions of work conducted and cost sharing are detailed in Supplemental Text (Additional File 1), which contains citations of references 34-39 and Additional Files 16, 17, 18, 19, 20 and is essentially an expanded version of Meth- ods. Additional material Additional file 1 Supplemental Text Click here for file [http://www.biomedcentral.com/content/supplementary/1471- 2164-10-582-S1.DOC] Additional file 2 Figure S1. SNP yield. The near-linear relationship between the number of SNPs and the product of the number of EST sequences for pairwise gen- otype comparisons is shown by plotting all values versus a linear regression line. Each axis is on a logarithmic scale. Oval shapes indicate a compari- son involving the wild barley accession OHU602. See text for additional details. Click here for file [http://www.biomedcentral.com/content/supplementary/1471- 2164-10-582-S2.DOC] Additional file 3 Table S1. All individual and consensus maps, including SxM bin mark- ers. Click here for file [http://www.biomedcentral.com/content/supplementary/1471- 2164-10-582-S3.XLS] Page 11 of 13 (page number not for citation purposes)