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Published November 10, 1989 | public
Journal Article

Genomic Sequencing and Methylation Analysis by Ligation Mediated PCR


Genomic sequencing permits studies of in vivo DNA methylation and protein-DNA interactions, but its use has been limited because of the complexity of the mammalian genome. A newly developed genomic sequencing procedure in which a ligation mediated polymerase chain reaction (PCR) is used generates high quality, reproducible sequence ladders starting with only 1 microgram of uncloned mammalian DNA per reaction. Different sequence ladders can be created simultaneously by inclusion of multiple primers and visualized separately by rehybridization. Relatively little radioactivity is needed for hybridization and exposure times are short. Methylation patterns in genomic DNA are readily detectable; for example, 17 CpG dinucleotides in the 5' region of human X-linked PGK-1 (phosphoglycerate kinase 1) were found to be methylated on an inactive human X chromosome, but unmethylated on an active X chromosome.

Additional Information

© 1989 American Association for the Advancement of Science. 11 July 1989; accepted2 6 September 1989. Supported by National Institute of Aging grant AG08196 (A.D.R.), NIH grants RR07003 and GM355262BW (B.W.), and a fellowship from the Deutsche Forschungsgemeinschaft (Pf212/1-1) (G-P.P.).

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