Construction of libraries for methylation sites by in-gel competitive reassociation (IGCR).
The in-gel competitive reassociation (IGCR) procedure was successfully applied to construct a comprehensive library enriched in DNA fragments containing C5mCGG sequences from mouse liver and brain genomic DNA. For IGCR, methylation-insensitive restriction enzyme (Msp I) digests were used as target DNA and methylation-sensitive restriction enzyme (Hpa II) digests as competitor DNA. Southern blot analysis indicated that 60 to 70% of the clones in the library were derived from the methylated sites and overall enrichment was 200- to 1000-fold. IGCR was further applied to construct a library for the sites differentially methylated between brain and liver DNA. In the library, approximately 20% of the Hpa II sites exhibited different degrees of methylation between these tissues.
Mizuguchi
R
R
Institute of Molecular and Cellular Biosciences, University of Tokyo, Japan.
Inoue
S
S
Kiyama
R
R
Oishi
M
M
D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000378:metabolism; D003001:Cloning, Molecular / Q000379:methods; D004247:DNA / Q000235:genetics*; D053938:DNA Fragmentation; D019175:DNA Methylation; D015723:Gene Library; D016678:Genome; D008099:Liver / Q000378:metabolism; D051379:Mice
D004247:DNA