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Genomic N(6)-methyladenine determination by MEKC with LIF

Krais, Annette M LU ; Cornelius, Michael G and Schmeiser, Heinz H. (2010) In Electrophoresis 31(21). p.51-3548
Abstract

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 oligodeoxyribonucleotide 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 ∼1... (More)

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 oligodeoxyribonucleotide 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 ∼1 N(6)mdA per 10(4) normal nucleotides (0.01%) using 1 μg of DNA as the matrix. After calibration with completely dam methylated λ DNA, the assay was applied to the analysis of various DNAs.

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published
keywords
Adenine, Animals, Boron Compounds, Cattle, Chromatography, Micellar Electrokinetic Capillary, DNA, DNA, Bacterial, DNA, Viral, Electrophoresis, Agar Gel, Ethylenediamines, Humans, Lasers, Sensitivity and Specificity, Spectrometry, Fluorescence, Journal Article, Research Support, Non-U.S. Gov't
in
Electrophoresis
volume
31
issue
21
pages
4 pages
publisher
John Wiley & Sons
external identifiers
  • scopus:77958605397
ISSN
0173-0835
DOI
10.1002/elps.201000357
language
English
LU publication?
no
id
a3302651-c9ac-4768-b7dd-eb6473905137
date added to LUP
2017-10-17 15:31:59
date last changed
2018-07-15 04:45:03
@article{a3302651-c9ac-4768-b7dd-eb6473905137,
  abstract     = {<p>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 oligodeoxyribonucleotide 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 ∼1 N(6)mdA per 10(4) normal nucleotides (0.01%) using 1 μg of DNA as the matrix. After calibration with completely dam methylated λ DNA, the assay was applied to the analysis of various DNAs.</p>},
  author       = {Krais, Annette M and Cornelius, Michael G and Schmeiser, Heinz H.},
  issn         = {0173-0835},
  keyword      = {Adenine,Animals,Boron Compounds,Cattle,Chromatography, Micellar Electrokinetic Capillary,DNA,DNA, Bacterial,DNA, Viral,Electrophoresis, Agar Gel,Ethylenediamines,Humans,Lasers,Sensitivity and Specificity,Spectrometry, Fluorescence,Journal Article,Research Support, Non-U.S. Gov't},
  language     = {eng},
  number       = {21},
  pages        = {51--3548},
  publisher    = {John Wiley & Sons},
  series       = {Electrophoresis},
  title        = {Genomic N(6)-methyladenine determination by MEKC with LIF},
  url          = {http://dx.doi.org/10.1002/elps.201000357},
  volume       = {31},
  year         = {2010},
}