Sensitive detection of oxidative DNA damage in cyanobacterial cells using supercoiling-sensitive quantitative PCR. (November 2018)

• Record Type: Journal Article
• Authors: Chen, Zhilan; Tian, Yun; Zhu, Chenhong; Liu, Biyun; Zhang, Yongyuan; Lu, Zhiying; Zhou, Qiaohong; Wu, Zhenbin
• Is Part Of: Chemosphere. Volume 211(2018)
• Page Start: 164
• Page End: 172
• Publication Date: 2018-11
• Subjects: Oxidative DNA damage -- Supercoiling-sensitive quantitative PCR -- Cyanobacterium -- Oxidative stress -- Exogenetic H2O2; Pollution -- Periodicals; Pollution -- Physiological effect -- Periodicals; Environmental sciences -- Periodicals; Atmospheric chemistry -- Periodicals; 551.511
• DOI: 10.1016/j.chemosphere.2018.06.154
• Languages: English
• ISSNs: 0045-6535

Abstract

Abstract: Supercoiling-sensitive quantitative PCR (ss-qPCR) is a sensitive technique to detect DNA damage in cultured animal cells and cultured/clinical human cells in vitro . In this study, we investigated whether the ss-qPCR method can be applied as a sensitive means to detect oxidative DNA damage in unicellular organisms. We used the model cyanobacterium Synechococcus elongatus PCC 7942 as a test organism and H2 O2 as an exogenetic oxidative toxicant. Results showed that a significant increase in the plasmid DNA damage of S. elongatus PCC 7942 was induced by H2 O2 in a dose- and time-dependent manner. The sensitivity of ss-qPCR in detecting DNA damage of the cyanobacterium was higher than the cell inhibition method (up to 255 times) as calculated from the slopes of fitted curves in the tested sub-toxic concentration range of 1–5 mM H2 O2 . Ss-qPCR also detected repairable low-intensity DNA damage in the cyanobacterium when DNA repair inhibitors were used. The detection limit of modified ss-qPCR was one tenth of that of previous methods. We also observed that ss-qPCR can be used to detect genomic DNA conformation change of cyanobacterium exposed to H2 O2 . Thus, this method will provide a powerful technical support for investigating the mechanisms of cyanobacterial DNA damage by environmental factors, especially intracellular reactive oxygen species enhancement-related factors. Graphical abstract: Image 1 Highlights: The ss-qPCR method can detect plasmid DNA damage in cyanobacterium. The plasmid DNA damage was in a dose- and time-dependent relationship with H2 O2 . The ss-qPCR method is more sensitive than the cell inhibition method (255 times). With DNA repair inhibitors, the detection limit can cut to one tenth of that. The ss-qPCR also can detect genomic DNA conformation change.