The cystic fibrosis transmembrane conductance regulator (CFTR) gene encodes a cAMP‐regulated chloride channel that is important in controlling the exchange of fluid and electrolytes across epithelial cells. Mutation of CFTR can lead to cystic fibrosis (CF), the most common lethal genetic disease in Caucasians. CF is a systemic illness with multiple organ systems affected including pulmonary, gastrointestinal, pancreatic, immune, endocrine, and reproductive systems. To understand the role of CFTR in the various tissues in which it is expressed, we generated a murine conditional null allele of Cftr (Cftr^fl10) in which loxP sites were inserted around exon 10 of the Cftr gene. The Cftr^fl10 allele was validated by generating constitutive Cftr null (Cftr^Δ10) mice using the protamine‐cre system. The Cftr^Δ10/Δ10 mice displayed almost identical phenotypes to previously published CF mouse models, including poor growth, decreased survival, intestinal obstruction, and loss of Cftr function as assessed by electrophysiology measurements on gut and nasal epithelium. Mice containing the conditional null Cftr allele will be useful in future studies to understand the role of Cftr in specific tissues and developmental time points and lead to a better understanding of CF disease. genesis 46:546–552, 2008. © 2008 Wiley‐Liss, Inc.