Open AccessConstruction of Libraries for Methylation Sites by In-gel Competitive Reassociation (IGCR)
Author(s) -
R Mizuguchi
Publication year - 1995
Publication title -
dna research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.647
H-Index - 98
eISSN - 1756-1663
pISSN - 1340-2838
DOI - 10.1093/dnares/2.5.219
Subject(s) - biology , dna methylation , restriction enzyme , dna , genomic dna , methylation , microbiology and biotechnology , southern blot , genomic library , genetics , gene , gene expression , base sequence
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.
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