Development of a KASP marker set for high-throughput genotyping in Japanese barley breeding programs with various end-use purposes.

Abstract

In barley (Hordeum vulgare L.), many DNA markers have been developed for the selection of traits related to various end-use purposes of breeding. To perform rapid marker-assisted selection of many lines, we developed Kompetitive Allele-Specific PCR (KASP) markers, which can be used for effective automatic genotyping of single nucleotide polymorphisms (SNPs). The KASP primers were designed for 17 SNPs in 14 genes related to important traits. The target allele of all primers tried was identified on the basis of high FAM fluorescence in comparison with that of HEX. To evaluate the suitability of the developed markers in breeding programs, we used them to genotype 62 representative cultivars and lines. Then, using six of the developed markers, we comprehensively analyzed a total of 2,941 lines collected from eight breeding sites with a genotyping success rate of 95.1%-99.8% (mean, 98.6%). All six markers showed differences in allele percentages among breeding programs, and specific allele combinations were observed in all four barley types. Our data will be useful for predicting phenotype segregation and designing cross combinations. The developed KASP marker set can be used for high-throughput genotyping and should make breeding more efficient when combined with an accelerated generation technique.