AbstractLoss‐of‐function approaches by the Cre/loxP technology have provided powerful tools for functional analyses of genes of interest expressed preferentially in a particular tissue. Here we describe the generation of transgenic mouse lines expressing Cre recombinase under the control of the promoter/enhancer unit of the gene for the α2 chain of collagen type I (Col1α2). As an expression vector, we used a P1‐derived artificial chromosome (PAC), which harbors ∼100 kb carrying the col1α2 gene. The improved coding sequence of the Cre recombinase was introduced to replace the first exon of col1α2. Cre expression was determined by immunohistochemistry and Cre‐mediated onset of β‐galactosidase expression in ROSA26R‐Cre reporter mice. In four analyzed transgenic lines, Cre recombinase was efficiently expressed during embryogenesis and in adult animals in cells of mesenchymal origin, such as dermal fibroblasts, mesenchymal cells of blood vessel walls, and cells in fibrous connective tissues surrounding internal organs. genesis 38:139–144, 2004. © 2004 Wiley‐Liss, Inc.