Simultaneous molecular karyotyping and mapping of viral DNA integration sites by 25-color COBRA-FISH

Combined binary ratio labeling (COBRA) fluorescence in situ hybridization ( FISH) allows 24-color FISH karyotyping of human metaphase chromosomes utili zing only four fluorochromes, instead of the five required for combinatoria l labeling procedures. Here we show that by introduction of a fifth fluoroc hrome, COBRA-FISH permits molecular cytogenetic mapping of viral integratio n sites in complex karyotypes in the context of a 24-color hybridization. W e were able to detect a single copy of the human papillomavirus 16 in the S iHa cell line and to confirm the site of integration at 13q21-31. We also d emonstrate the gene mapping possibility of 25-color hybridization by detect ing a MYC cosmid on normal metaphase chromosomes. the possibility of mappin g single-copy probes in the background of 24-color hybridization expands th e tools for cytogenetic mapping of DNA sequences and will contribute to the understanding of the role of viral integration and chromosome rearrangemen t in virus-mediated carcinogenesis. Genes Chromosomes Cancer 28:92-97, 2000 . (C) 2000 WiIey-Liss, Inc.