PCR-Based Analysis of Mitochondrial DNA Copy Number, Mitochondrial DNA Damage, and Nuclear DNA Damage.

Date
2016-02-01
Authors
Gonzalez-Hunt, Claudia P
Rooney, John P
Ryde, Ian T
Anbalagan, Charumathi
Joglekar, Rashmi
Meyer, Joel N
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Abstract
Because of the role that DNA damage and depletion play in human disease, it is important to develop and improve tools to assess these endpoints. This unit describes PCR-based methods to measure nuclear and mitochondrial DNA damage and copy number. Long amplicon quantitative polymerase chain reaction (LA-QPCR) is used to detect DNA damage by measuring the number of polymerase-inhibiting lesions present based on the amount of PCR amplification; real-time PCR (RT-PCR) is used to calculate genome content. In this unit, we provide step-by-step instructions to perform these assays in Homo sapiens, Mus musculus, Rattus norvegicus, Caenorhabditis elegans, Drosophila melanogaster, Danio rerio, Oryzias latipes, Fundulus grandis, and Fundulus heteroclitus, and discuss the advantages and disadvantages of these assays.
Type
Journal article
Subject
C. elegans
DNA damage
mitochondrial DNA
mitochondrial DNA copy number
qPCR
Animals
Cell Nucleus
DNA Copy Number Variations
DNA Damage
DNA Mutational Analysis
DNA Primers
DNA, Mitochondrial
Humans
Polymerase Chain Reaction
Permalink
https://hdl.handle.net/10161/12422
Published Version (Please cite this version)
10.1002/0471140856.tx2011s67
Publication Info
Gonzalez-Hunt, Claudia P; Rooney, John P; Ryde, Ian T; Anbalagan, Charumathi; Joglekar, Rashmi; & Meyer, Joel N (2016). PCR-Based Analysis of Mitochondrial DNA Copy Number, Mitochondrial DNA Damage, and Nuclear DNA Damage. Curr Protoc Toxicol, 67. pp. 20.11.1-20.11.25. 10.1002/0471140856.tx2011s67. Retrieved from https://hdl.handle.net/10161/12422.
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Scholars@Duke

Gonzalez-Hunt

Claudia Gonzalez-Hunt

Research Associate, Senior
Research Interests: Mitochondrial biology, genome integrity, Parkinson's disease, neurodegeneration
Meyer

Joel Meyer

Associate Professor of Environmental Genomics in the Division of Environmental Sciences and Policy
Dr. Meyer studies the effects of toxic agents and stressors on human and wildlife health. He is particularly interested in understanding the mechanisms by which environmental agents cause DNA damage, the molecular processes that organisms employ to protect prevent and repair DNA damage, and genetic differences that may lead to increased or decreased sensitivity to DNA damage. Mitochondrial DNA damage and repair, as well as mitochondrial function in general, are a particular focus. He studies
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