Shimizu Hiroaki, Ishikawa Goro, Aoki Hideyuki, Nakata Masaru, Tanaka Junichi
Kyushu Okinawa Agricultural Research Center, National Agriculture and Food Research Organization (NARO), Izumi 496, Chikugo, Fukuoka 833-0041, Japan.
Institute of Crop Science, NARO, 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8518, Japan.
Breed Sci. 2025 Apr;75(2):129-138. doi: 10.1270/jsbbs.24052. Epub 2025 Apr 2.
In barley ( 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.
在大麦（L.）中，已经开发了许多DNA标记用于选择与育种的各种最终用途相关的性状。为了对多个品系进行快速的标记辅助选择，我们开发了竞争性等位基因特异性PCR（KASP）标记，其可用于单核苷酸多态性（SNP）的有效自动基因分型。针对与重要性状相关的14个基因中的17个SNP设计了KASP引物。根据与HEX相比更高的FAM荧光来鉴定所有测试引物的目标等位基因。为了评估所开发标记在育种计划中的适用性，我们用它们对62个代表性品种和品系进行基因分型。然后，使用六个开发的标记，我们对从八个育种地点收集的总共2941个品系进行了综合分析，基因分型成功率为95.1%-99.8%（平均98.6%）。所有六个标记在不同育种计划中的等位基因百分比均存在差异，并且在所有四种大麦类型中均观察到特定的等位基因组合。我们的数据将有助于预测表型分离和设计杂交组合。所开发的KASP标记集可用于高通量基因分型，并且与加速世代技术结合使用时应能提高育种效率。
