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)
Watanabe
S
S
Shionogi Research Laboratories, Osaka, Japan.
Kawai
J
J
Hirotsune
S
S
Suzuki
H
H
Hirose
K
K
Taga
C
C
Ozawa
N
N
Fushiki
S
S
Hayashizaki
Y
Y
D000818:Animals; D001921:Brain / Q000254:growth & development / Q000378:metabolism*; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D015252:Deoxyribonucleases, Type II Site-Specific; D004586:Electrophoresis / Q000379:methods*; D005821:Genetic Techniques; D016678:Genome; D008745:Methylation; D051379:Mice; D008809:Mice, Inbred C3H; D009928:Organ Specificity; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism
D012333:RNA, Messenger; D004247:DNA; D015252:Deoxyribonucleases, Type II Site-Specific; C038565:GCGGCCGC-specific type II deoxyribonucleases