Transferring multiple genes into the same cell allows for the combination of genetic correction, marking, selection and conditional elimination of transduced cells or the reconstitution of multisubunit components and synergistic pathways. However, this cannot be reliably accomplished by current gene transfer technologies. Based on the finding that some cellular promoters intrinsically promote divergent transcription, we have developed synthetic bidirectional promoters that mediate coordinate transcription of two mRNAs in a ubiquitous or a tissue-specific manner. Lentiviral vectors incorporating the new promoters enabled efficient dual gene transfer in several tissues in vivo after direct delivery or transgenesis, and in a human gene therapy model. Because divergent gene pairs, likely transcribed from shared promoters, are common in the genome, the synthetic promoters that we developed may mimic a well-represented feature of transcription. Vectors incorporating these promoters should increase the power of gene function studies and expand the reach and safety of gene therapy.