Abstract
BackgroundOne of the key goals of oak genomics research is to identify genes of adaptive significance. This information may help to improve the conservation of adaptive genetic variation and the management of forests to increase their health and productivity. Deep-coverage large-insert genomic libraries are a crucial tool for attaining this objective. We report herein the construction of a BAC library for Quercus robur, its characterization and an analysis of BAC end sequences.ResultsThe EcoRI library generated consisted of 92,160 clones, 7% of which had no insert. Levels of chloroplast and mitochondrial contamination were below 3% and 1%, respectively. Mean clone insert size was estimated at 135 kb. The library represents 12 haploid genome equivalents and, the likelihood of finding a particular oak sequence of interest is greater than 99%. Genome coverage was confirmed by PCR screening of the library with 60 unique genetic loci sampled from the genetic linkage map. In total, about 20,000 high-quality BAC end sequences (BESs) were generated by sequencing 15,000 clones. Roughly 5.88% of the combined BAC end sequence length corresponded to known retroelements while ab initio repeat detection methods identified 41 additional repeats. Collectively, characterized and novel repeats account for roughly 8.94% of the genome. Further analysis of the BESs revealed 1,823 putative genes suggesting at least 29,340 genes in the oak genome. BESs were aligned with the genome sequences of Arabidopsis thaliana, Vitis vinifera and Populus trichocarpa. One putative collinear microsyntenic region encoding an alcohol acyl transferase protein was observed between oak and chromosome 2 of V. vinifera.ConclusionsThis BAC library provides a new resource for genomic studies, including SSR marker development, physical mapping, comparative genomics and genome sequencing. BES analysis provided insight into the structure of the oak genome. These sequences will be used in the assembly of a future genome sequence for oak.
Highlights
One of the key goals of oak genomics research is to identify genes of adaptive significance
BAC library characterization Estimation of mean insert size This library consists of 92,160 clones stored into 240 384-well plates
We evaluated the mean size of BAC inserts by randomly selecting 189 clones, extracting their DNA and digesting it with the rare cutter enzyme NotI for analysis by PFGE (Figure 1A)
Summary
One of the key goals of oak genomics research is to identify genes of adaptive significance. Large collections of oak expressed sequence tags (ESTs) have been generated from various tissues and developmental stages, including 130,000 Sanger sequences and 2 M 454-reads, available from public databases [5]. This catalog constitutes a useful resource for detecting candidate genes controlling traits of interest and for the development of new genetic markers for forward genetics approaches (linkage mapping and QTL detection, association mapping) for dissection of the genetic architecture of adaptive traits [6,7,8,9]. Little is known about the overall structure of the oak genome
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