We developed alloplasmic (cytoplasm substitution) lines of 14 Japanese bread wheat cultivars with Aegilops mutica cytoplasm to examine the effects of cytoplasm on agronomic characters. Compared with euplasmic wheat having normal cytoplasm, the alloplasmic lines generally showed increased spikelet number per spike but decreased floret number per spikelet, resulting in the decreased grain number per spike. However the grain number per spike varied depending on genotype; in the alloplasmic lines of Nanbukomugi, Nebarigoshi and Fukusayaka, no decrease of the grain number occurred. All alloplasmic lines showed delayed heading time (5 to 15 days in the field), and the degree of heading delay depended on genotype. In the spring wheat cultivars such as Chikugoizumi, the degree of heading delay due to the alien cytoplasm is large, while it is small in the winter wheat cultivars such as Haruibuki. The alloplasmic lines exhibited the lower expression levels of flowering promoter gene VERNALIZATION 1 (VRN1) compared with the euplasmic wheat. In the spring wheat cultivars, the difference in the VRN1 expression levels between the alloplasmic lines and euplasmic lines are large, leading to the significant delay in heading time. The alloplasmic lines may be useful for the development of varieties adapted to global warming.