RESTRICTION LANDMARK GENOMIC SCANNING METHOD AND ITS VARIOUS APPLICATIONS

We have developed a new genome scanning method (restriction landmark g enomic scanning (RLGS), based on the new concept of using restriction enzyme sites as landmarks. RLGS employs direct end labeling of the gen omic DNA digested with a restriction enzyme and two-dimensional electr ophoresis with high-resolution. Its advantages are: (i) high-speed sca nning ability, allowing simultaneous scanning of thousands of restrict ion landmarks; (ii) extension of the scanning field using different ki nds of landmarks in an additional series of electrophoresis; (iii) app lication to any type of organism because of direct-labeling of restric tion enzyme sites and no hybridization procedure; and (iv) reflection of the copy number of the restriction landmark by the spot intensity w hich enables distinction of haploid and diploid genomic DNAs. The RLG S method has various applications because it can be used to scan for p hysical genomic DNA states, such as amplification, deletion and methyl ation. The copy number of the locus of a restriction landmark can be e stimated by the spot intensity to find either an amplified or deleted region. The methylation state of genomic DNA can also be discovered by use of a methylation-sensitive restriction enzyme sites as a restrict ion landmark (restriction landmark genomic scanning for screening meth ylated sites, RLGS-M). This article introduces the basic principle of RLGS and its applications to the analysis of cancer, mouse mutant DNAs and tissue-specific methylation, showing the usefulness of RLGS for a variety of biological fields.