A new technique for determining the distribution of N7-methylguanine using an automated DNA sequencer. Carcinogenesis 1991 Nov;12(11):2089-92
Date
11/01/1991Pubmed ID
1682064DOI
10.1093/carcin/12.11.2089Scopus ID
2-s2.0-0026001896 (requires institutional sign-in at Scopus site) 5 CitationsAbstract
We have developed a method to determine rapidly the sequence specificity of DNA alkylation resulting from chemical treatment. The utility of this approach is demonstrated here in a study of the sequence specificity of alkylation by dimethylsulphate (DMS). The method is independent of the sequence chosen and makes use of the polymerase chain reaction (PCR) to generate a fluorescently labelled DNA target. In this study, a 302 bp segment of the Escherichia coli lacI gene was amplified and the product purified by liquid chromatography on a Mono Q column. This DNA was alkylated with DMS and treated with hot piperidine to produce single-strand breaks at sites of N7 alkylation. The distribution of the break points, and hence the position and extent of alkylation, were determined on an Applied Biosystems 370A automated DNA sequencer.
Author List
Shoukry S, Anderson MW, Glickman BWMESH terms used to index this publication - Major topics in bold
AlkylationDNA
DNA Damage
Escherichia coli
Gene Amplification
Guanine
Lac Operon
Molecular Sequence Data
Oligonucleotides
Piperidines
Polymerase Chain Reaction
Polymorphism, Restriction Fragment Length
Sulfuric Acid Esters