Global gene deletion studies in mice and humans have established the pivotal role of runt related transcription factor‐2 (Runx2) in both intramembranous and endochondral ossification processes during skeletogenesis. In this study, we for the first time generated mice carrying a conditional Runx2 allele with exon 4, which encodes the Runt domain, flanked by loxP sites. These mice were crossed with α1(I)‐collagen‐Cre or α1(II)‐collagen‐Cre transgenic mice to obtain osteoblast‐specific or chondrocyte‐specific Runx2 deficient mice, respectively. As seen in Runx2^−/− mice, perinatal lethality was observed in α1(II)‐Cre;Runx2^flox/flox mice, but this was not the case in animals in which α1(I)‐collagen‐Cre was used to delete Runx2. When using double‐staining with Alizarin red for mineralized matrix and Alcian blue for cartilaginous matrix, we observed previously that mineralization was totally absent at embryonic day 15.5 (E15.5) throughout the body in Runx2^−/− mice, but was found in areas undergoing intramembranous ossification such as skull and clavicles in α1(II)‐Cre;Runx2^flox/flox mice. In newborn α1(II)‐Cre;Runx2^flox/flox mice, mineralization impairment was restricted to skeletal areas undergoing endochondral ossification including long bones and vertebrae. In contrast, no apparent skeletal abnormalities were seen in mutant embryo, newborn, and 3‐week‐old to 6‐week old‐mice in which Runx2 had been deleted with the α1(I)‐collagen‐Cre driver. These results suggest that Runx2 is absolutely required for endochondral ossification during embryonic and postnatal skeletogenesis, but that disrupting its expression in already committed osteoblasts as achieved here with the α1(I)‐collagen‐Cre driver does not affect overtly intramembranous and endochondral ossification. The Runx2 floxed allele established here is undoubtedly useful for investigating the role of Runx2 in particular cells. © 2013 American Society for Bone and Mineral Research.