microarray-based genotyping and genetic mapping approach for highly heterozygous outcrossing species enables localization of a large fraction of the unassembled Populus trichocarpa genome sequence

• Published in: The Plant journal : for cell and molecular biology 2009-06, Vol.58 (6), p.1054-1067
• Main Authors: Drost, Derek R, Novaes, Evandro, Boaventura-Novaes, Carolina, Benedict, Catherine I, Brown, Ryan S, Yin, Tongming, Tuskan, Gerald A, Kirst, Matias
• Format: Article
• Language: English
• Subjects: Base Sequence; Biological and medical sciences; Botany; Chromosome Mapping - methods; Classical genetics, quantitative genetics, hybrids; DNA Probes; DNA, Plant - genetics; Fundamental and applied biological sciences. Psychology; Gene expression; gene expression marker; Genetics of eukaryotes. Biological and molecular evolution; genome assembly; Genome, Plant; Genomics; Genotype; Genotype & phenotype; Methods, theories and miscellaneous; microarray; Microsatellite Repeats; Molecular Sequence Data; Oligonucleotide Array Sequence Analysis - methods; Polymorphism, Genetic; Populus; Populus - genetics; Populus trichocarpa; Research methodology; single-feature polymorphism
• ISSN: 0960-7412; 1365-313X
• DOI: 10.1111/j.1365-313X.2009.03828.x

Microarrays have demonstrated significant power for genome-wide analyses of gene expression, and recently have also revolutionized the genetic analysis of segregating populations by genotyping thousands of loci in a single assay. Although microarray-based genotyping approaches have been successfully applied in yeast and several inbred plant species, their power has not been proven in an outcrossing species with extensive genetic diversity. Here we have developed methods for high-throughput microarray-based genotyping in such species using a pseudo-backcross progeny of 154 individuals of Populus trichocarpa and P. deltoides analyzed with long-oligonucleotide in situ-synthesized microarray probes. Our analysis resulted in high-confidence genotypes for 719 single-feature polymorphism (SFP) and 1014 gene expression marker (GEM) candidates. Using these genotypes and an established microsatellite (SSR) framework map, we produced a high-density genetic map comprising over 600 SFPs, GEMs and SSRs. The abundance of gene-based markers allowed us to localize over 35 million base pairs of previously unplaced whole-genome shotgun (WGS) scaffold sequence to putative locations in the genome of P. trichocarpa. A high proportion of sampled scaffolds could be verified for their placement with independently mapped SSRs, demonstrating the previously un-utilized power that high-density genotyping can provide in the context of map-based WGS sequence reassembly. Our results provide a substantial contribution to the continued improvement of the Populus genome assembly, while demonstrating the feasibility of microarray-based genotyping in a highly heterozygous population. The strategies presented are applicable to genetic mapping efforts in all plant species with similarly high levels of genetic diversity.