BarPlex v1.0: a multiplex PCR-based enrichment of genome-wide short segments that enable genetic studies in barley

Abstract Over the past 15 years sequencing methodologies have advanced greatly enabling high-throughput sequencing based genotyping of crop plants. In this study, we developed BarPlex v1.0, a robust and cost-efficient genotyping approach in barley ( Hordeum vulgare L.). In this multiplex PCR-based amplification of five-hundred genome-wide segments, followed by high-throughput sequencing of barcoded PCR products, we obtained hundreds to thousands of polymorphic markers. Comparison of genotyping with BarPlex v1.0 to genotyping-by-sequencing (GBS) revealed a similar genetic diversity. The polymorphic markers revealed by BarPlex v1.0 were highly accurate, with an average sequencing depth >700x and a data missing rate <0.5%. By analyzing 1,068 genotypes of wild barley (brittle rachis; Hordeum vulgare ssp. spontaneum L.), Tibetan semi-wild barley (brittle rachis), landraces, cultivars, as well as an F 2 population, this assay has been robust in studies of population diversity, variety pedigree, heterozygosity discrimination, linkage mapping, as well as genome-wide association study (GWAS). Notably, a diversity analysis in a population of Tibetan semi-wild barley suggested a close relationship with Chinese landraces, but a dramatic decrease in its genetic diversity, inferring that Tibet was not a center of domestication for the native wild barley.