The influence of the semidwarfing gene sd1 derived from the rice cultivar Jukkoku (Jukkoku_sd1) and IR8 (IR8_sd1), which contributed to the Green Revolution, d60 from Hokuriku 100, as well as the combination of sd1 and d60 (Jukkoku_sd1 plus d60 and IR8_sd1 plus d60), was investigated using isogenic lines raised by backcrossing with the cultivar Koshihikari. The isogenic lines carrying Jukkoku_sd1, IR8_sd1, d60, Jukkoku_sd1 plus d60, and IR8_sd1 plus d60 had considerably shorter culm lengths than Koshihikari by 19.2%, 22.8%, 26.0%, 45.1%, and 43.4%, respectively. The sd1 plus d60 lines showed additively reduced culms, indicating that the function of d60 was different from sd1. In contrast to the culm reduction, Jukkoku_sd1 showed productive merit with a panicle length of 2.5% greater than the origin. MiSeq next-generation sequencer was used to optimize a minimum scale to detect Jukkoku_sd1 in practical breeding. Mapping with the reference genome of Nipponbare gained the average depths of Koshihikari Jukkoku_sd1 and Koshihikari being 9.17 and 7.29, respectively. Comparing the vcf files of the entire genomes of Koshihikari Jukkoku_sd1 and the virtual Koshihikari revealed a G to T SNP at position 38,382,746 in the sd1 locus on chromosome 1 of Koshihikari, causing a loss-of-function mutation of GA20-oxidase.