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Microassay for DNA methyltransferase.

R L Adams1, A Rinaldi, C Seivwright

  • 1Department of Biochemistry, University of Glasgow, UK.

Journal of Biochemical and Biophysical Methods
|January 1, 1991
PubMed
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A new microassay detects DNA methylase activity in just 50 cells. This sensitive method uses [3H]AdoMet and synthetic DNA for efficient DNA methyltransferase detection.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Background:

  • DNA methylases play crucial roles in gene regulation and cellular processes.
  • Sensitive detection methods are needed for studying DNA methylase activity in limited cell samples.

Purpose of the Study:

  • To develop and describe a novel microassay for quantifying DNA methylase activity.
  • To demonstrate the assay's sensitivity using a small number of cells.

Main Methods:

  • A microassay was developed utilizing lysed tissue culture cells.
  • Incubation with high specific activity [3H]AdoMet (S-adenosylmethionine) and synthetic DNA substrate poly[d(I-C).d(I-C)].
  • Assay includes ribonuclease and product DNA isolation via phenol extraction after protease digestion.

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Main Results:

  • The microassay can detect DNA methylase activity in as few as 50 tissue culture cells.
  • The assay is sensitive and requires minimal starting material.

Conclusions:

  • A highly sensitive microassay for DNA methylase activity has been established.
  • This method is suitable for analyzing DNA methylase activity in small cell populations.