Submucosal glands (SMGs) are the major site of expression of the cystic fibrosis (CF) transmembrane conductance regulator gene (CFTR) in the human lung. As such, SMGs may be a critical component of CF lung disease pathogenesis and an important target for gene therapy. Gene-targeted mouse models exist for CF and these are used to validate gene therapy or other interventions and to dissect CF phenotypes. It is important, therefore, to compare human and mouse SMGs. We show that SMGs in the mouse are similar in structure, cell types, and Cftr expression to those in the human. Murine SMGs were found to be present in the proximal regions of the trachea at the same density as in humans but, unlike in humans, did not extend below the trachea. Upon investigation of homozygous Cftr tm1HGU and Cftr tm1G551D mutant mice, SMGs were found to extend more distally than those in wild-type control mice (P < 0.05). To investigate the development of SMGs we generated aggregation chimeric mice. Chimeric offspring contained a contribution of transgenic cells that were detectable either by DNA in situ hybridization (reiterated beta-globin transgene TgN[Hbb-bl]83Clo) or beta-galactosidase histochemistry (Lac Z reporter gene TgR[ROSA26]- 26Sor). Analysis of the distribution of transgenic cells in chimeric SMGs suggests that SMGs are clonally derived.