Interspecific gene flow (introgression) is an important source of new genetic variation, but selection against it can reinforce reproductive barriers between interbreeding species. We used an experimental approach to trace the role of chromosomal inversions and incompatibility genes in preventing introgression between two partly sympatric Drosophila virilis group species, D. flavomontana and D. montana. We backcrossed F₁ hybrid females from a cross between D. flavomontana female and D. montana male with the males of the parental species for two generations and sequenced pools of parental strains and their reciprocal 2^nd generation backcross (BC₂mon and BC₂fla) females. Contrasting the observed amount of introgression (mean hybrid index, HI) in BC₂ female pools along the genome to simulations under different scenarios allowed us to identify chromosomal regions of restricted and increased introgression. We found no deviation from the HI expected under a neutral null model for any chromosome for the BC₂mon pool, suggesting no evidence for genetic incompatibilities in backcrosses towards D. montana. In contrast, the BC₂fla pool showed high variation in the observed HI between different chromosomes, and massive reduction of introgression on the X chromosome (large X-effect). We find that this observation is compatible with reduced recombination combined with at least one dominant incompatibility locus residing within the X inversion(s). Overall, our study suggests that genetic incompatibilities arising within chromosomal inversions can play an important role in speciation.