The Effects of Sub-Inhibitory Levels of Chloramphenicol on pBR322 Plasmid Copy Number in Escherichia coli DH5α Cells

Abstract

While humans use antibiotics at inhibitory levels to prevent microbial growth, antibiotics are normally found at sub-inhibitory concentrations in nature. The role and effects of sub-inhibitory levels of antibiotics is one of the key areas of research in microbial science today. In this experiment, the effects of sub-inhibitory levels of chloramphenicol on pBR322 plasmid copy number in Escherichia coli DH5 α cells were examined. E. coli DH5 α cells with pBR322 plasmid were grown in different concentrations of chloramphenicol ranging from 1 to 5 µg/ml. The cells were enzymatically lysed and the DNA extracted through a phenol:chloroform:isoamyl alcohol extraction. The total bacterial DNA was run on a 0.8% agarose gel; using the ratio of genomic DNA to pBR322 DNA, the plasmid copy numbers per genome were calculated. It was found that 1 µg/ml chloramphenicol quantities had no significant affect when compared to the control; however, 3-5 µg/ml of chloramphenicol caused pBR322 concentrations to double. Due to assay insensitivity, there was no significant difference between the 3 µg/ml and 5 µg/ml samples.