Accessibility to tissue-specific genes from methylation profiles of mouse brain genomic DNA

The DNA methylation status of a large number of genomic loci is visualized simultaneously and quantitatively as two-dimensional gel spots in the newly developed restriction landmark genomic scanning with a methylation-sensitive restriction enzyme (RLGS-M). Here, we demonstrate that RLGS-M using NorI as a methylation-sensitive enzyme could also scan gene loci of mammalian genomes, since almost all of the NotI loci corresponding to randomly chosen RLGS-M spots were located near or in transcriptional units (6 out of 7 NotI-linking clones) when mouse brain genomic DNA was used. This supports the previous prediction that most NotI sites are located in CpG islands (Lindsay and Bird, Nature 1987, 327, 336-338). Furthermore, beginning with RLGS-M spots we examined how to approach their corresponding RNA messages, whose expression may be associated with methylation. We compared RLGS-M patterns among various developmental stages of the mouse brain from embryonic day 9.5 to postnatal 8 weeks or among in vitro cell lines, and detected alterations of RLGS-M spots which were due to methylation of NotI sites. Two experiments using NotI-linking clones or polymerase chain reaction (PCR) were carried out to approach to their corresponding RNA messages. Consequently, we isolated two PCR-amplified clones (# 15 and # 91) which corresponded to methylatable loci and gave positive signals to mRNA from the adult brain. Furthermore, we identified two NotI-linking clones (C211 and C198) whose corresponding NotI loci localized near or at transcriptional units and were methylated in cell lines.(ABSTRACT TRUNCATED AT 250 WORDS)