In the last 20 years, we have observed an exponential growth of the DNA sequence data and simular increase in the volume of DNA polymorphism data generated by numerous molecular marker technologies. Most of the investment, and therefore progress, concentrated on human genome and genomes of selected model species. Diversity Arrays Technology (DArT), developed over a decade ago, was among the first "democratizing" genotyping technologies, as its performance was primarily driven by the level of DNA sequence variation in the species rather than by the level of financial investment. DArT also proved more robust to genome size and ploidy-level differences among approximately 60 organisms for which DArT was developed to date compared to other high-throughput genotyping technologies. The success of DArT in a number of organisms, including a wide range of "orphan crops," can be attributed to the simplicity of underlying concepts: DArT combines genome complexity reduction methods enriching for genic regions with a highly parallel assay readout on a number of "open-access" microarray platforms. The quantitative nature of the assay enabled a number of applications in which allelic frequencies can be estimated from DArT arrays. A typical DArT assay tests for polymorphism tens of thousands of genomic loci with the final number of markers reported (hundreds to thousands) reflecting the level of DNA sequence variation in the tested loci. Detailed DArT methods, protocols, and a range of their application examples as well as DArT's evolution path are presented.

Diversity Arrays Technology (DArT) can detect and type DNA variation at several hundred genomic loci in parallel without relying on sequence information. Here we show that it can be effectively applied to genetic mapping and diversity analyses of barley, a species with a 5,000-Mbp genome. We tested several complexity reduction methods and selected two that generated the most polymorphic genomic representations. Arrays containing individual fragments from these representations generated DArT fingerprints with a genotype call rate of 98.0% and a scoring reproducibility of at least 99. A lthough 50 years have passed since the structure of DNA was deciphered (1), the study of DNA variation emerged as a field of scientific endeavor only in the last 25 years. Two groups of technologies were developing in parallel from the very beginning: DNA sequencing and molecular markers. DNA sequencing technology developed quickly from proof of concept (2, 3) to an automated process (4), enabling the field of genomics. Molecular marker technologies progressed rapidly as well. Based on the Southern blot technique (5), Botstein et al. (6) developed the restriction fragment length polymorphism (RFLP) technique as a method for creating genetic linkage maps.Development of the PCR technique spawned two important molecular marker techniques: amplified fragment length polymorphism (AFLP) (7) and simple sequence repeats (8). Thousands of studies using molecular markers in plants, including hundreds in barley, have been published but are not referenced because of space limitations.DNA sequencing and molecular marker technologies started to merge when the accumulated sequence data began to yield information on sequence variation among different accessions of the same species. It was soon noted that single-nucleotide polymorphism (SNP) is the most abundant marker type, promising nearly unlimited supply of markers (9). Many alternatives were developed for the SNP assay (primer extension, selective ligation) and the platform to type assays in high throughput (DNA chip, printed and self-assembling arrays, matrix-assisted laser desorption ionization͞ time-of-flight mass spectroscopy) (10)(11)(12)(13)(14).For humans and a limited number of model organisms, the throughput of SNP assays has increased impressively, and assay costs have decreased correspondingly. Yet discovering sequence polymorphism in nonmodel species is difficult, which is particularly true for many crops with limited resources and often complex, polyploid genomes. We have developed Diversity Arrays Technology (DArT) to enable whole-genome profiling of such crops without the need of sequence information. DArT is based on microarray hybridizations that detect the presence versus absence of individual fragments in genomic representations as described by Jaccoud et al. (15).For our initial proof-of-concept study, we selected a species with a simple genome (rice) and used AFLP-like complexity reduction methods to generate genomic representations (15). Here we apply a non-AFLP version ...

Despite a substantial investment in the development of panels of single nucleotide polymorphism (SNP) markers, the simple sequence repeat (SSR) technology with a limited multiplexing capability remains a standard, even for applications requiring whole-genome information. Diversity arrays technology (DArT) types hundreds to thousands of genomic loci in parallel, as previously demonstrated in a number diploid plant species. Here we show that DArT performs similarly well for the hexaploid genome of bread wheat (Triticum aestivum L.). The methodology previously used to generate DArT fingerprints of barley also generated a large number of high-quality markers in wheat (99.8% allele-calling concordance and approximately 95% call rate). The genetic relationships among bread wheat cultivars revealed by DArT coincided with knowledge generated with other methods, and even closely related cultivars could be distinguished. To verify the Mendelian behaviour of DArT markers, we typed a set of 90 Cranbrook x Halberd doubled haploid lines for which a framework (FW) map comprising a total of 339 SSR, restriction fragment length polymorphism (RFLP) and amplified fragment length polymorphism (AFLP) markers was available. We added an equal number of DArT markers to this data set and also incorporated 71 sequence tagged microsatellite (STM) markers. A comparison of logarithm of the odds (LOD) scores, call rates and the degree of genome coverage indicated that the quality and information content of the DArT data set was comparable to that of the combined SSR/RFLP/AFLP data set of the FW map.

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