Abstract: [Objective]Chromosome substitution lines between Chinese spring and Synthetic 6x under different water treatments were used to locate the gene controlling spikelet progress and drought tolerance.[Method]A series of spikelet differentiation progress were observed.[Result] The result indicated that spikelet progress of various substitution lines and that of parents, including spike and flower anlage periods, were almost alike. Under normal irrigation,2B substitution line is the latest in the progress of spikelet differentiation, 7D substitution line is the second, while Chinese spring and other substitution lines are the rapidest. The spikelet differentiation progress of 2D substitution line is much slower than Chinese spring and other substitution lines under drought condition.[Conclusin] It is concluded that the genes inhibiting spikelet differentiation may be located on 2B and 7D chromosome, and that the genes of drought tolerance may be located on 2D chromosome. The results can provide theoretic basis for the genetic and breeding research concerned with drought resistance and water saving.