Preparation of hydrogels for the study of the effects of spatial confinement on DNA

Abstract

Model studies under biomimetic nondilute conditions are important to understand the effects of a cellular environment on nucleic acid molecules. Hydrogels forming a network structure have been used as a model for investigating the effect of a confined molecular environment on proteins but not on nucleic acids. The present study developed a method for preparing agarose and polyacrylamide gels for measuring the thermal stability of a double-stranded DNA polymer poly(dA)/poly(dT) in a confined environment. Gel materials were prepared by the addition of DNA before the formation of the gel network, but experiments for the analysis of DNA melting curves had several limitations. The shape and extent of hyperchromicity of the melting curve of poly(dA)/poly(dT) embedded in the agarose gel were similar to those obtained with the buffer solution. On the other hand, the UV absorbance of the polyacrylamide gel largely increased after heating, and thermal melting of poly(dA)/poly(dT) in the polyacrylamide gel started at a high absorbance, giving ambiguous results. Hydrogels prepared according to the procedure developed in this study would be useful for developing new strategies for studying the effects of confinement on nucleic acid molecules.