DNA polymerase profiling

Summerer, Daniel

DNA polymerase profiling

Files

Summerer.pdf(27.82 MB)

Date

2008

Authors

Summerer, Daniel

Series

Methods in Molecular Biology

Publication type

Journal article

Published in

Methods in molecular biology (Clifton, N.J.) ; 429 (2008). - pp. 225-235. - ISSN 1064-3745

Abstract

We report a simple homogeneous fluorescence assay for quantification of DNA polymerase function in high throughput. The fluorescence signal is generated by the DNA polymerase triggering opening of a molecular beacon extension of the template strand. A resulting distance alteration is reported by fluorescence resonance energy transfer between two dyes introduced into the molecular beacon stem. We describe real-time reaction profiling of two model DNA polymerases. We demonstrate kinetic characterization, rapid optimization of reaction conditions, and inhibitor profiling using the presented assay. Furthermore, to supersede purification steps in screening procedures of DNA polymerase mutant libraries, detection of enzymatic activity in bacterial expression lysates is described.

Subject (DDC)

540 Chemistry

Cite This

ISO 690

SUMMERER, Daniel, 2008. DNA polymerase profiling. In: Methods in molecular biology (Clifton, N.J.). 429, pp. 225-235. ISSN 1064-3745. Available under: doi: 10.1007/978-1-60327-040-3_16

BibTex

@article{Summerer2008polym-353,
  year={2008},
  doi={10.1007/978-1-60327-040-3_16},
  title={DNA polymerase profiling},
  volume={429},
  issn={1064-3745},
  journal={Methods in molecular biology (Clifton, N.J.)},
  pages={225--235},
  author={Summerer, Daniel}
}

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