Determination of methylation specificity of sequence-specific DNA methyltransferases using matrix assisted laser desorption/ionization time-of-flight mass spectrometry

Takashi Tamura, Yoshinori Araki, Seiji Yamaoka, Kenji Inagaki, Hidehiko Tanaka

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

We describe here a sensitive and straightforward method for characterizing the methylation specificity of type II DNA methyltransferase (MTase) using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. DNA substrate, prepared by ligation of a commercially available oligonucleotide, was modified by the subject MTase, and was derivatized to a mixture of single-stranded oligonucleotides through endonuclease treatment, heat-denaturation and limited digestion by 3'-terminus-specific phosphodiesterase I. MALDI-TOF mass spectrometry was used to determine the mass differences between the digestion products, and the methylated nucleotide was explicitly identified by the mass increase of 14 Da due to the base modification. The method was applicable to the three representative MTases M.EcoRI, M.BamHI and M.HaeIII.

Original languageEnglish
Pages (from-to)4162-4164
Number of pages3
JournalNucleic acids research.
Volume25
Issue number20
DOIs
Publication statusPublished - Oct 15 1997

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Methyltransferases
Oligonucleotides
Methylation
Digestion
Phosphatidyl-N-Methylethanolamine N-Methyltransferase
Mass Spectrometry
Lasers
Phosphodiesterase I
Endonucleases
DNA
Ligation
Nucleotides
Hot Temperature
DNA modification methylase EcoRI

ASJC Scopus subject areas

  • Genetics

Cite this

Determination of methylation specificity of sequence-specific DNA methyltransferases using matrix assisted laser desorption/ionization time-of-flight mass spectrometry. / Tamura, Takashi; Araki, Yoshinori; Yamaoka, Seiji; Inagaki, Kenji; Tanaka, Hidehiko.

In: Nucleic acids research., Vol. 25, No. 20, 15.10.1997, p. 4162-4164.

Research output: Contribution to journalArticle

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