Primary chondrocyte isolation and culture is a useful tool to characterize how cellular perturbations impact chondrocyte behavior and mineralization in vitro. This protocol conveys methods for isolating and culturing primary chondrocytes from costal and growth plate cartilage. Following gross dissection of the neonatal murine anterior rib cage or long bone growth plate cartilage, chondrocytes are isolated via enzymatic digestion and plated at high density. Genetic perturbation of plated primary murine chondrocytes using viral infection of Cre recombinase to excise floxed alleles and/or overexpress genes of interest are also described.