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Issue 5, 2012
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Exonucleolytic degradation of high-density labeled DNA studied by fluorescence correlation spectroscopy

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Abstract

The exonucleolytic degradation of high-density labeled DNA by exonuclease III was monitored using two-color fluorescence correlation spectroscopy (FCS). One strand of the double stranded template DNA was labeled on either one or two base types and additionally at one end via a 5′ Cy5 tagged primer. Exonucleolytic degradation was followed via the diffusion time, the brightness of the remaining DNA as well as the concentration of released labeled bases. We found a hydrolyzation rate of about 11 to 17 nucleotides per minute per enzyme (nt/min/enzyme) for high-density labeled DNA, which is by a factor of about 4 slower than for unlabeled DNA. The exonucleolytic degradation of a 488 base pair long double stranded DNA resulted in a short double stranded DNA segment of 112 ± 40 base pairs (bp) length with two single-stranded tails.

Graphical abstract: Exonucleolytic degradation of high-density labeled DNA studied by fluorescence correlation spectroscopy

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Publication details

The article was received on 20 Sep 2011, accepted on 05 Jan 2012 and first published on 23 Jan 2012


Article type: Paper
DOI: 10.1039/C2AN15879E
Citation: Analyst, 2012,137, 1160-1167
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    Exonucleolytic degradation of high-density labeled DNA studied by fluorescence correlation spectroscopy

    N. Ehrlich, K. Anhalt, H. Paulsen, S. Brakmann and C. G. Hübner, Analyst, 2012, 137, 1160
    DOI: 10.1039/C2AN15879E

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