Preharvest sprouting (PHS), caused by rain after maturity, leads to a severe degradation of buckwheat quality. We previously reported the PHS tolerance of several natural resources and landraces in Japan, and have bred PHS-tolerant lines. Here, we investigate the rates of PHS in our breeding lines and leading cultivars in Japan. The new lines ‘Kyukei 28’ and ‘Kyukei 29’ had better PHS tolerance than all leading cultivars. To reveal the PHS tolerance inherited by these lines, we performed genetic analysis using crosses between these lines and a self-compatible line, ‘Kyukei SC7’. The rate of germination in F₂ segregating lines showed different segregation patterns, suggesting that major tolerance genes in ‘Kyukei 28’ (KY28) are recessive and those in ‘Kyukei 29’ (KY29) are dominant. This information could help to accelerate the breeding of buckwheat lines with PHS tolerance and develop DNA markers for the selection of tolerant lines.