This unit describes an important advance in fiber-FISH technology called molecular combing, in which single DNA molecules are bound by one or both ends to a surface and stretched in a uniform and parallel manner by a receding meniscus. This technique is gentle on the molecules, rapid, and easy to perform. Reliable, quantitative information for genome-wide studies can be obtained without the need for other techniques and a large number of accurate measurements can be made in a single experiment. The authors provide detailed protocols for basic molecular combing, high-resolution physical mapping, and gene-dosage approaches as well as support protocols outlining surface preparation, DNA solution preparation, and probe labeling.