Understanding the role of manganese in the prevention of DNA damage within E. coli helps us understand the survival of bacteria to the human immune response and has the potential to aid in the development of novel antibiotics. Manganese is a beneficial cofactor for proteins in E. coli as it does not participate in the chemistry that forms free radicals that iron does. The manganese transport protein MntH had previously been knocked out from E. coli reporter strains CSH102 and CSH104 to inhibit their ability to intake manganese. The presence of manganese had been shown to decrease the rates of oxidative mutations in E. coli reporter strain CSH104. In this work, a Comet assay was used to try and visualize direct DNA damage levels in these E. coli strains. The comet assay is a gel electrophoresis technique used to measure and visualize DNA strand fragmentation within an individual cell. Comet assays were performed on cells from CSH104 and CSH104Δ with and without hydrogen peroxide induced DNA damage. Tails indicating fragmented DNA were visualized and compared between the parent and knockout strains. It was found that unlike mammalian control cells, E. coli required harsher lysis conditions to visualize DNA.

COMET-based assay for detecting DNA damage in Mn(II) deficient strains of E. coli

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