A genomic scanning method for higher organisms using restriction sites as landmarks

Abstract

We have developed a powerful genomic scanning method, termed "restriction landmark genomic scanning," that is useful for analysis of the genomic DNA of higher organisms using restriction sites as landmarks. Genomic DNA is radioactively labeled at cleavage sites specific for a rare cleaving restriction enzyme and then size-fractionated in one dimension. The fractionated DNA is further digested with another more frequently occurring enzyme and separated in the second dimension. This procedure gives a two-dimensional pattern with thousands of scattered spots corresponding to sites for the first enzyme, indicating that the genome of mammals can be scanned at approximately 1-megabase intervals. The position and intensity of a spot reflect its locus and the copy number of the corresponding restriction site, respectively, based on the nature of the end-labeling system. Therefore, this method is widely applicable to genome mapping or detection of alterations in a genome.