Gene transcription is regulated by the formation of protein-DNA complexes that influence the rate of specific initiation of transcription by RNA polymerase. Recent experimental advances allowing the identification of cis regulatory sequences that specify the binding of trans acting protein factors have made significant contributions to our understanding of the mechanistic complexities of transcriptional regulation. These methodologies have prompted the use of similar strategies to elucidate transcriptional control mechanisms involved in the tissue specific and developmental regulation of pulmonary surfactant protein gene expression. The purpose of this review is to describe various methodologies by which molecular biologists identify and subsequently assay regions of nucleic acids presumed to be integral in gene regulation at the level of transcription. It is well established that genes encoding surfactant proteins are subject to regulation by hormones, cytokines, and a variety of biologically active reagents. Perhaps future studies utilizing molecular tools outlined in this review will be valuable in identification of DNA sequences and protein factors required for the regulation of lung surfactant genes.