By using a CaCO₃ co-precipitation technique, p53 expression plasmids and doxorubicin hydrochloride (DOX) were encapsulated in nano-sized CaCO₃/DNA/DOX co-precipitates for co-delivery of genes and drugs. Under a certain Ca²⁺/CO₃²⁻ ratio in the co-precipitation, both plasmid DNA and drugs could be loaded in the CaCO₃/DNA/DOX nanoparticles with high encapsulation efficiency. The in vitrocell inhibition of the CaCO₃/DNA/DOX nanoparticles was evaluated in HeLa cells by a MTT assay. The results showed the simultaneous treatment by gene and drug could induce cell apoptosis and completely inhibit the cell proliferation. The CaCO₃/DNA/DOX nanoparticles exhibited a high cell inhibition rate of about 75%, indicating that the CaCO₃/DNA/DOX nanoparticles could effectively mediate gene transfection and deliver the drug to the cells. Compared with the gene delivery system (CaCO₃/DNA nanoparticles) or the free drug DOX, the co-delivery system (CaCO₃/DNA/DOX nanoparticles) exhibits enhanced cell inhibition rate. The calcium carbonate based approach has great potential in the preparation of gene and drug co-delivery systems, and the CaCO₃/DNA/DOX nanoparticles have promising applications in cancer treatment.