Viscoelastic characterization of single-stranded DNA from Escherichia coli

Abstract

Single-stranded DNA released from E. coli wild type and mutant cells by alkaline-EDTA-detergent was analyzed using the recently developed biophysical technique of viscoelastometry. Under the lysis conditions used, it was possible to detect single strands of molecular weight approximately 2 times 10-9 daltons. Little difference was detected in the size of single-stranded DNA from log phase vs. stationary phase cultures, or from cells treated with chloramphenicol to allow completion of replicating chromosomes. The largest single strands from ligase overproducing, endonuclease minus, and pol A1 mutants were likewise of approximately the same size as wild type, but were present in smaller yields. The reduction in single-strand molecular weight as a result of heating intact cells was investigated as a function of time and temperature. Heating at 37 degrees C for up to 20 min produced no additional single-strand breaks, but temperatures from 45 to 65 degrees introduced breaks. Solutions maintained at pH 12.5 were not stable indefinitely, and the relative viscosity of such solutions was found to decrease over a period of several hours.