Genomic N6-methyladenine determination by MEKC with LIF

2'-Deoxy-N-6-methyladenosine (N(6)mdA) is frequently found in prokaryotic and unicellular eukaryotic genomes. Although methylated bases represent only a minor fraction of the genome, they, however, exhibit strong biological effects. Here, we report a fast and sensitive method for the quantification of global adenine methylation in DNA. The method is based on a recently developed procedure consisting of fluorescence labeling of deoxyribonucleotides with BODIPY FL EDA and analysis by CE with LIF. An oligodeoxy-ribonucleotide site specifically modified with N(6)mdA was used for peak assignment, to establish separation conditions and to determine the LOD. The method yielded a LOD for N(6)mdA of 280 pM (1.4 amol), which is equivalent to similar to 1 N(6)mdA per 10(4) normal nucleotides (0.01%) using 1 mu g of DNA as the matrix. After calibration with completely dam methylated lambda DNA, the assay was applied to the analysis of various DNAs.