A new mapping system, based on nonisotopic in situ hybridization combined with fluorescent staining of replicated prometaphase R-bands, is described. Replication of the bands is achieved by treatment of thymidine-synchronized cells with bromodeoxyuridine. The human COL2A1 gene was mapped to band 12q13.11-q13.12 in this manner, to illustrate the potential of the technique for improving the precision of chromosomal mapping and physical ordering of genes.