Restriction Landmark Genomic Scanning
RLGS is a two-dimensional gel electrophoresis method that is used as a genome scanning technique to survey genomes for copy number changes as well as changes in DNA methylation status of promoter sequences.
RLGS was developed as a high-speed genome scanning technique for the identification of DNA amplification as well as DNA methylation changes. For RLGS, genomic DNA is restriction digested with a landmark enzyme, usually a rare-cutting enzyme (e.g., NotI). The restriction ends are end labeled in a fill-in reaction using [α-32P]-labeled nucleotides. In the following second restriction digest (e.g., EcoRV), the DNA fragments are cut into smaller fragments suitable for separation in an agarose disk gel electrophoresis (first dimension). The next step is an in-gel restriction digestion with a third enzyme (e.g., HinfI) that generates fragments with a size range between 100 and 2,000 base pairs. DNA fragments are separated in a polyacrylamide gel (second...