Using second- or third-chromosome substitution lines of Drosophila melanogaster, the genetic variation of inducibility and amylase specific activities in three media (starch, normal and glucose) were investigated. Genetic factors on both the second and third chromosomes were responsible for the variation in amylase specific activity and inducibility. In glucose medium, the genetic variance of amylase specific activity estimated for the second-chromosome substitution lines was larger than that for the third-chromosome substitution lines; however, for starch medium and inducibility, the variance was larger for the third-chromosome substitution lines. High correlations for the second-chromosome substitution lines and low correlations for the third-chromosome substitution lines were observed for amylase specific activities in different media. These results suggest that the genetic factor(s) responsible for inducibility or amylase activity variation in an induced medium such as starch should be on the third chromosome and those in the non-induced medium such as glucose should be on the second chromosome. The functional roles of the factors on the second and third chromosomes would be the repression and induction of amylase, respectively.