We have constructed a high-resolution cytogenetic map of human chromosome 2 with 434 newly isolated cosmid markers by means of direct R-banding fluorescence in situ hybridization. Two markers were mapped to the centromeric region, 173 to the short arm, and 259 to the long arm. The clones were evenly distributed along the entire chromosome, although a tendency toward clustering in R-positive bands was observed. The mapped cosmids provide useful landmarks for construction of a contig map and for positional cloning of cancer-associated genes, as well as genes responsible for hereditary diseases.