COMET ASSAY TO STUDY DNA DAMAGE CAUSED BY DIFFERENT STRESS INDUCERS IN THE RIBOFLAVIN OVERPRODUCER EREMOTHECIUM ASHBYII

Abstract

Objectives: The present study focuses on investigating morphological changes and genotoxicity effects due to physical and chemical mutagens such as riboflavin, ethidium bromide (EtBr), pH, and ultraviolet (UV).

Methods: Riboflavin doses were given at 0.2 mM. 0.4 mM, 0.8 mM, and 1.0 mM concentration, pH stress doses were given by adjusting the pH of the medium to pH 3.0, pH 5.0, and pH 7.0 whereas EtBr stress doses were 1.0 μL and 0.5 μL and UV radiation stress doses were given for 5 min, 10 min, 15 min. Our study reports at using a modified alkaline yeast comet assay methodology to detect DNA damages in Eremothecium ashbyii caused by stress inducers. Fluorescence microscopy was used to detect the comets and CaspLab software to quantify the intensity of DNA damages.

Results and Conclusion: The metrics employed for the quantification of DNA damage under the specified stressful circumstances were, tail DNA, olive tail moment (OTM), and the tail moment (TM). The statistical analysis of the DNA damage doses with CaspLab parameters to detect DNA damage shows maximum DNA damages at pH 5.0 with the highest peak for OTM value followed by UV damage at 15 min. Our study reports the first case using the comet assay technique to detect the DNA damages in the filamentous E. ashbyii cells.