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A modified and automated version of the 'Fluorimetric Detection of Alkaline DNA Unwinding' method to quantify formation and repair of DNA strand breaks

A modified and automated version of the 'Fluorimetric Detection of Alkaline DNA Unwinding' method to quantify formation and repair of DNA strand breaks

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MORENO-VILLANUEVA, María, Ragen PFEIFFER, Thilo SINDLINGER, Alan LEAKE, Marcus MÜLLER, Thomas B. L. KIRKWOOD, Alexander BÜRKLE, 2009. A modified and automated version of the 'Fluorimetric Detection of Alkaline DNA Unwinding' method to quantify formation and repair of DNA strand breaks. In: BMC Biotechnology. 9(April), 39. ISSN 1472-6750. eISSN 1472-6750. Available under: doi: 10.1186/1472-6750-9-39

@article{MorenoVillanueva2009modif-6790, title={A modified and automated version of the 'Fluorimetric Detection of Alkaline DNA Unwinding' method to quantify formation and repair of DNA strand breaks}, year={2009}, doi={10.1186/1472-6750-9-39}, number={April}, volume={9}, issn={1472-6750}, journal={BMC Biotechnology}, author={Moreno-Villanueva, María and Pfeiffer, Ragen and Sindlinger, Thilo and Leake, Alan and Müller, Marcus and Kirkwood, Thomas B. L. and Bürkle, Alexander}, note={Article Number: 39} }

deposit-license Kirkwood, Thomas B. L. Leake, Alan Sindlinger, Thilo Bürkle, Alexander 2011-03-24T17:29:12Z First publ. in: BMC Biotechnology ; 9 (2009), April. - 39 Pfeiffer, Ragen Müller, Marcus 2011-03-24T17:29:12Z Moreno-Villanueva, María Müller, Marcus Pfeiffer, Ragen 2009 Moreno-Villanueva, María Sindlinger, Thilo Bürkle, Alexander A modified and automated version of the 'Fluorimetric Detection of Alkaline DNA Unwinding' method to quantify formation and repair of DNA strand breaks application/pdf Background: Formation and repair of DNA single-strand breaks are important parameters in the assessment of DNA damage and repair occurring in live cells. The 'Fluorimetric Detection of Alkaline DNA Unwinding (FADU)' method [Birnboim HC, Jevcak JJ. Cancer Res (1981) 41:1889 1892] is a sensitive procedure to quantify DNA strand breaks, yet it is very tedious to perform.<br />Results: In order (i) to render the FADU assay more convenient and robust, (ii) to increase throughput, and (iii) to reduce the number of cells needed, we have established a modified assay version that is largely automated and is based on the use of a liquid handling device. The assay is operated in a 96-well format, thus greatly increasing throughput. The number of cells required has been reduced to less than 10,000 per data point. The threshold for detection of X-ray-induced DNA strand breaks is 0.13 Gy. The total assay time required for a typical experiment to assess DNA strand break repair is 4 5 hours.<br />Conclusion: We have established a robust and convenient method measuring of formation and repair of DNA single-strand breaks in live cells. While the sensitivity of our method is comparable to current assays, throughput is massively increased while operator time is decreased. Kirkwood, Thomas B. L. eng Leake, Alan

Dateiabrufe seit 01.10.2014 (Informationen über die Zugriffsstatistik)

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