Genetic basis of reproductive isolation is an important issue to study speciation. Hybrid incompatibility (HI) in the forms of hybrid lethality or sterility, are commonly found in closely related species causing reproductive isolation and thus lead to speciation. One classical model for studying HI is the hybrids from the cross ofDrosophila melanogaster and D. simulans. The cross between D. melanogaster females and D. simulans males only produces sterile F1 females and hybrid males are larval lethal. The reciprocal cross yields only sterile F1 males while the hybrid females are embryonic lethal. Using attached-X chromosome strain, it is proposed that the hybrid female lethality is caused by the present of D. melanogaster X chromosome (Xm) and maternal product from D. simulans. To identify loci which contribute to the hybrid lethality, we conduct a genetic screen by crossing D. simulans females to males of the D. melanogaster X chromosome duplication lines that contain an extra X fragments on Y chromosomes in Zhr background. We have screened 22 lines covering 97.22 % of the X chromosome, and the results show that 2 of the 22 duplications have significant effect on hybrid male lethality. Our results indicate that there are at least two regions on the Xm chromosome contributing to HI between D. melanogaster and D. simulans.