title: Adapting Genotyping-by-Sequencing for Rice F2 Populations.
abstract: Rapid and cost-effective genotyping of large mapping populations can be achieved by sequencing a reduced representation of the genome of every individual in a given population, and using that information to generate genetic markers. A customized genotyping-by-sequencing (GBS) pipeline was developed to genotype a rice F2 population from a cross ofOryza sativassp.japonicacv. Nipponbare and the African wild rice speciesO. longistaminataWhile most GBS pipelines aim to analyze mainly homozygous populations, we attempted to genotype a highly heterozygous F2 population. We show how species- and population-specific improvements of established protocols can drastically increase sample throughput and genotype quality. Using as few as 50,000 reads for some individuals (134,000 reads on average), we were able to generate up to 8154 informative SNP markers in 1081 F2 individuals. Additionally, the effects of enzyme choice, read coverage, and data postprocessing are evaluated. Using GBS-derived markers, we were able to assemble a genetic map of 1536 cM. To demonstrate the usefulness of our GBS pipeline, we determined quantitative trait loci (QTL) for the number of tillers. We were able to map four QTL to chromosomes 1, 3, 4, and 8, and partially confirm their effects using introgression lines. We provide an example of how to successfully use GBS with heterozygous F2 populations. By using the comparatively low-cost MiSeq platform, we show that the GBS method is flexible and cost-effective, even for smaller laboratories.
authors.affiliation: Bioscience and Biotechnology Center, Nagoya University, 464-8601, Japan.
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Furuta
authors.affiliation: Bioscience and Biotechnology Center, Nagoya University, 464-8601, Japan.
authors.forename: Motoyuki
authors.initials: M
authors.lastname: Ashikari
authors.affiliation: Plant Breeding Division, International Rice Research Institute, 1301 Manila, Philippines.
authors.forename: Kshirod K
authors.initials: KK
authors.lastname: Jena
authors.affiliation: Associated Field Science and Research Center, Nagoya University, 470-0151, Japan kdoi@agr.nagoya-u.ac.jp reuscher@agr.nagoya-u.ac.jp.
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Doi
authors.affiliation: Bioscience and Biotechnology Center, Nagoya University, 464-8601, Japan kdoi@agr.nagoya-u.ac.jp reuscher@agr.nagoya-u.ac.jp.
authors.forename: Stefan
authors.initials: S
authors.lastname: Reuscher
mesh_terms: D000483:Alleles; D032461:Chromosomes, Plant / Q000235:genetics; D003433:Crosses, Genetic; D005787:Gene Frequency / Q000235:genetics; D005819:Genetic Markers; D060005:Genotyping Techniques; D007178:Inbreeding; D012275:Oryza / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D040641:Quantitative Trait Loci / Q000235:genetics; D017422:Sequence Analysis, DNA
chemical_list: D005819:Genetic Markers
keywords: SNP marker; genotyping-by-sequencing; rice breeding; trait mapping