PUBLICATION
Simple and fast quantification of DNA damage by real-time PCR, and its application to nuclear and mitochondrial DNA from multiple tissues of aging zebrafish
- Authors
- Zhu, S., Coffman, J.A.
- ID
- ZDB-PUB-170712-11
- Date
- 2017
- Source
- BMC research notes 10: 269 (Journal)
- Registered Authors
- Keywords
- Aging, DNA damage, Mitochondrial, Nuclear, Quantitative PCR, Zebrafish
- MeSH Terms
-
- Age Factors
- Animals
- Brain/metabolism
- Cell Nucleus/genetics*
- DNA Damage*
- DNA, Mitochondrial*
- Heart
- Liver/metabolism
- Muscle, Skeletal/metabolism
- Real-Time Polymerase Chain Reaction/methods*
- Zebrafish/genetics*
- PubMed
- 28693618 Full text @ BMC Res. Notes
Citation
Zhu, S., Coffman, J.A. (2017) Simple and fast quantification of DNA damage by real-time PCR, and its application to nuclear and mitochondrial DNA from multiple tissues of aging zebrafish. BMC research notes. 10:269.
Abstract
We describe a real-time (rt) PCR-based method of quantifying DNA damage, adapted from the long-run rtPCR method of DNA damage quantification (LORD-Q) developed by Lehle et al. (Nucleic Acids Res 42(6):e41, 2014). We show that semi-long run rtPCR, which generates amplicons half the length of those generated in LORD-Q, provides equivalent sensitivity for detecting low lesion frequencies, and better sensitivity for detecting high frequencies. The smaller amplicon size greatly facilitates PCR optimization and allows greater flexibility in the use of detection dyes, and a modified data analysis method simplifies the calculation of lesion frequency. The method was used to measure DNA damage in the nuclear and mitochondrial genomes of different tissues in zebrafish of different ages. We find that nuclear DNA damage generally increases with age, and that the amount of mitochondrial DNA damage varies substantially between tissues, increasing with age in liver and brain but not in heart or skeletal muscle, the latter having the highest levels of damage irrespective of age.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping