Five Drosophila melanogaster strains showing high interspecific crossability with D. simulans males, derived from the previous screening of a set of autosomal plwB transposants, were selected and the effect of the plwB insertion under the same genetic background (white strain) on the interspecific crossability was examined. This phenotype was recessive in the two strains but semidominant in the other three strains. Trans-heterozygotes, however, showed high interspecific crossability compared with the parental homozygotes, suggesting some epistatic interactions between them. In three strains, the effect of the plwB insertion region in different backgrounds (w1118 strain) on the crossability was also tested. Homozygotes of a strain (#687) showed high interspecific crossability comparable to the w background, while homozygotes of both #68 and #783 strains showed lower crossability than w1118. Although #783 heterozygotes showed intermediate values between #783 homozygotes and w1118, #68 heterozygotes showed a significantly higher insemination rate than w1118 and the #68 homozygotes. These results suggest that the region around the plwB insertion sites of #68 and #783 affects the interspecific crossability either positively or negatively depending on the genetic background. In all the stocks, positive correlation between interspecific crossability and the intraspecific mating speed was detected.