Development of Kompetitive Allele Specific PCR Markers for Anaerobic Germination 1 Locus in Rice

Abstract

The anaerobic germination 1 (AG1) locus, which confers tolerance to the anaerobic germination of rice seed, has been previously identified and a gel-based InDel DNA marker developed for marker-assisted selection (MAS). However, there is a need for marker development for high-throughput genotyping in order to increase the breeding efficiency. Single nucleotide polymorphisms (SNPs)-based Kompetitive Allele Specific PCR (KASP) markers enable high-accuracy and high-throughput analyses for the genotyping of AG1 locus. In the present study, we developed one AG1 InDel-specific and four flanking KASP markers, which can be used as the foreground and recombination markers, respectively, for the AG1 locus. Of those, three KASP markers developed were validated with Dasan-AG1 (BC3F2) segregating lines; the AG1 InDel-specific KASP marker was 100% consistent with the existing AG1 gel-based marker, while the two flanking markers showed 70.3 and 66.7 percent consistency to the AG1 gel-based marker, respectively. In addition, the five KASP markers developed were further used to genotype 78 Korean and 95 foreign rice varieties. The genotyping results with 172 varieties revealed that most of the Korean varieties possessed the AG1 locus; however, a wide variation was observed in the foreign varieties for AG1 locus. Hence, the KASP markers developed for AG1 locus show promise as useful tools for expediting the breeding of varieties tolerant to anaerobic germination stress via high-throughput genotyping.