Extremely low frequency electromagnetic field (ELF-EMF) exposure is attracting increased attention as a possible disease-inducing factor. The in vivo effects of short-term and long-term ELF-EMF exposure on male Drosophila melanogaster were studied using transcriptomic analysis for preliminary screening and QRT-PCR for further verification. Transcriptomic analysis indicated that 439 genes were up-regulated and 874 genes were down-regulated following short-term exposures and that 514 genes were up-regulated and 1206 genes were down-regulated following long-term exposures (expression >2- or <0.5-fold, respectively). In addition, there are 238 up-regulated genes and 598 down-regulated genes in the intersection of short-term and long-term exposure (expression >2- or <0.5-fold). The DEGs (differentially expressed genes) in D. melanogaster following short-term exposures were involved in metabolic processes, cytoskeletal organization, mitotic spindle organization, cell death, protein modification and proteolysis. Long-term exposure let to changes in expression of genes involved in metabolic processes, response to stress, mitotic spindle organization, aging, cell death and cellular respiration. In the intersection of short-term and long-term exposure, a series of DEGs were related to apoptosis, aging, immunological stress and reproduction. To check the ELF-EMF effects on reproduction, some experiments on male reproduction ability were performed. Their results indicated that short-term ELF-EMF exposure may decrease the reproductive ability of males, but long-term exposures had no effect on reproductive ability. Down-regulation of ark gene in the exposed males suggests that the decrease in reproductive capacity may be induced by the effects of ELF-EMF exposure on spermatogenesis through the caspase pathway. QRT-PCR analysis confirmed that jra, ark and decay genes were down regulated in males exposed for 1 Generation (1G) and 72 h, which suggests that apoptosis may be inhibited in vivo. ELF-EMF exposure may have accelerated cell senescence, as suggested by the down-regulation of both cat and jra genes and the up-regulation of hsp22 gene. Up-regulation of totA and hsp22 genes during exposure suggests that exposed flies might induce an in vivo immune response to counter the adverse effects encountered during ELF-EMF exposure. Down-regulation of cat genes suggests that the partial oxidative protection system might be restrained, especially during short-term exposures. This study demonstrates the bioeffects of ELF-EMF exposure and provides evidence for understanding the in vivo mechanisms of ELF-EMF exposure on male D. melanogaster.