Abstract
BackgroundA number of molecular marker technologies have allowed important advances in the understanding of the genetics and evolution of Eucalyptus, a genus that includes over 700 species, some of which are used worldwide in plantation forestry. Nevertheless, the average marker density achieved with current technologies remains at the level of a few hundred markers per population. Furthermore, the transferability of markers produced with most existing technology across species and pedigrees is usually very limited. High throughput, combined with wide genome coverage and high transferability are necessary to increase the resolution, speed and utility of molecular marker technology in eucalypts. We report the development of a high-density DArT genome profiling resource and demonstrate its potential for genome-wide diversity analysis and linkage mapping in several species of Eucalyptus.FindingsAfter testing several genome complexity reduction methods we identified the PstI/TaqI method as the most effective for Eucalyptus and developed 18 genomic libraries from PstI/TaqI representations of 64 different Eucalyptus species. A total of 23,808 cloned DNA fragments were screened and 13,300 (56%) were found to be polymorphic among 284 individuals. After a redundancy analysis, 6,528 markers were selected for the operational array and these were supplemented with 1,152 additional clones taken from a library made from the E. grandis tree whose genome has been sequenced. Performance validation for diversity studies revealed 4,752 polymorphic markers among 174 individuals. Additionally, 5,013 markers showed segregation when screened using six inter-specific mapping pedigrees, with an average of 2,211 polymorphic markers per pedigree and a minimum of 859 polymorphic markers that were shared between any two pedigrees.ConclusionsThis operational DArT array will deliver 1,000-2,000 polymorphic markers for linkage mapping in most eucalypt pedigrees and thus provide high genome coverage. This array will also provide a high-throughput platform for population genetics and phylogenetics in Eucalyptus. The transferability of DArT across species and pedigrees is particularly valuable for a large genus such as Eucalyptus and will facilitate the transfer of information between different studies. Furthermore, the DArT marker array will provide a high-resolution link between phenotypes in populations and the Eucalyptus reference genome, which will soon be completed.
Highlights
A number of molecular marker technologies have allowed important advances in the understanding of the genetics and evolution of Eucalyptus, a genus that includes over 700 species, some of which are used worldwide in plantation forestry
This operational Diversity Arrays Technology (DArT) array will deliver 1,000-2,000 polymorphic markers for linkage mapping in most eucalypt pedigrees and provide high genome coverage
The transferability of DArT across species and pedigrees is valuable for a large genus such as Eucalyptus and will facilitate the transfer of information between different studies
Summary
A number of molecular marker technologies have allowed important advances in the understanding of the genetics and evolution of Eucalyptus, a genus that includes over 700 species, some of which are used worldwide in plantation forestry. High throughput, combined with wide genome coverage and high transferability are necessary to increase the resolution, speed and utility of molecular marker technology in eucalypts. A number of molecular marker technologies have been developed and used for species of Eucalyptus in the last 20 years [1] Each of these technologies allowed important advances in the understanding of the multifaceted genetics, evolution and breeding of this vast genus that sample and because AFLP is gel-based it is relatively labour-intensive. We report the development of the first version of a high density operational DArT genotyping microarray with over 7,000 markers and demonstrate its potential for diversity and linkage mapping studies in species of Eucalyptus across the two most important subgenera
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