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Quantitative, Real-time Analysis of Base Excision Repair Activity in Cell Lysates Utilizing Lesion-specific Molecular Beacons
 

Quantitative, Real-time Analysis of Base Excision Repair Activity in Cell Lysates Utilizing Lesion-specific Molecular Beacons

Article DOI: 10.3791/4168-v • 15:01 min • August 6th, 2012
August 6th, 2012 •
1,2, 3, 1,2,4
1Department of Pharmacology & Chemical Biology, University of Pittsburgh School of Medicine, 2Hillman Cancer Center, University of Pittsburgh Cancer Institute, 3Department of Experimental Therapy, The Netherlands Cancer Institute, 4Department of Human Genetics, University of Pittsburgh School of Public Health

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We describe a method for the quantitative, real-time measurement of DNA glycosylase and AP endonuclease activities in cell nuclear lysates. The assay yields rates of DNA Repair activity amenable to kinetic analysis and is adaptable for quantification of DNA Repair activity in tissue and tumor lysates or with purified proteins.

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Quantitative Analysis Real-time Measurement Base Excision Repair DNA Glycosylase AP Endonuclease Molecular Beacons Substrate Deoxyoligonucleotide Carboxyfluorescein (6-FAM) Dabcyl Stem-loop Structure Nucleotides Base Pairs Förster Resonance Energy Transfer (FRET) Fluorescence DNA Repair Fluorescence Values Real-time Assessment PCR Instruments
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