Electrorelease of Escherichia coli nucleoids

Abstract

Bacterial chromosome is assembled and folded into one or several nucleoids, depending on the metabolic status of the cell. Development of reliable nucleoid isolation protocols has always been an objective for researchers. A rapid and reproducible procedure for isolation of E. coli nucleoids is described here, while the cell envelope is maintained. Membrane dispersions and vesicles were prepared by lysozyme-EDTA treatment with subsequent rupture of the spheroplasts by electric field. Under these conditions the yield of electroreleased nucleoids was around 90%. The extent of DNA-envelope contacts was determined by light microscopy employing phase contrast and fluorescence modes.