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dc.contributor.authorEvans, Stephen Owenen_NZ
dc.contributor.authorJameson, Michael B.en_NZ
dc.contributor.authorCursons, Raymond T.en_NZ
dc.contributor.authorPeters, Linda M.en_NZ
dc.contributor.authorBird, Steveen_NZ
dc.contributor.authorJacobson, Gregory M.en_NZ
dc.date.accessioned2017-01-16T00:39:31Z
dc.date.available2016en_NZ
dc.date.available2017-01-16T00:39:31Z
dc.date.issued2016en_NZ
dc.identifier.citationEvans, S. O., Jameson, M. B., Cursons, R. T., Peters, L. M., Bird, S., & Jacobson, G. M. (2016). Development of a qPCR method to measure mitochondrial and genomic DNA damage with application to chemotherapy-induced DNA damage and cryopreserved cells. Biology, 5(4). http://doi.org/10.3390/biology5040039en
dc.identifier.urihttps://hdl.handle.net/10289/10841
dc.description.abstractDNA damage quantitation assays such as the comet assay have focused on the measurement of total nuclear damage per cell. The adoption of PCR-based techniques to quantify DNA damage has enabled sequence- and organelle-specific assessment of DNA lesions. Here we report on an adaptation of a qPCR technique to assess DNA damage in nuclear and mitochondrial targets relative to control. Novel aspects of this assay include application of the assay to the Rotor-Gene platform with optimized DNA polymerase/fluorophore/primer set combination in a touchdown PCR protocol. Assay validation was performed using ultraviolet C radiation in A549 and THP1 cancer cell lines. A comparison was made to the comet assay applied to peripheral blood mononuclear cells, and an estimation of the effects of cryopreservation on ultraviolet C-induced DNA damage was carried out. Finally, dose responses for DNA damage were measured in peripheral blood mononuclear cells following exposure to the cytotoxic agents bleomycin and cisplatin. We show reproducible experimental outputs across the tested conditions and concordance with published findings with respect to mitochondrial and nuclear genotoxic susceptibilities. The application of this DNA damage assay to a wide range of clinical and laboratory-derived samples is both feasible and resource-efficient.en_NZ
dc.format.mimetypeapplication/pdf
dc.language.isoen
dc.publisherMDPI AGen_NZ
dc.relation.urihttp://www.mdpi.com/2079-7737/5/4/39en_NZ
dc.rightsThis is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).
dc.subjectLORD-Qen_NZ
dc.subjectqPCRen_NZ
dc.subjectcomet assayen_NZ
dc.subjectDNA damageen_NZ
dc.subjectPBMCen_NZ
dc.subjectTHP1en_NZ
dc.subjectA549en_NZ
dc.subjectbleomycinen_NZ
dc.subjectcisplatinen_NZ
dc.titleDevelopment of a qPCR method to measure mitochondrial and genomic DNA damage with application to chemotherapy-induced DNA damage and cryopreserved cellsen_NZ
dc.typeJournal Article
dc.identifier.doi10.3390/biology5040039en_NZ
dc.relation.isPartOfBiologyen_NZ
pubs.elements-id143384
pubs.issue4en_NZ
pubs.volume5en_NZ
dc.identifier.eissn2079-7737en_NZ


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