Fluorescence spectral characteristics of novel asymmetric monomethine cyanine dyes in nucleic acid solutions.

Six new asymmetric monomethine cyanine dyes have been synthesized and their fluorescence characteristics in the presence of nucleic acids studied. The new dyes have no fluorescence of their own in water solutions upon excitation at 480 nm but they become strongly fluorescent in the presence of nucleic acids. The fluorescence maxima of the investigated dyes are found at 525-545 nm when bound to dsDNA and around 600 nm upon binding to RNA and ssDNA. Fluorescence quenching studies with increasing concentrations of NaCl indicate that the cyanine dyes have a mixed (intercalating and groove binding) type of interaction with dsDNA.

Heat shock and osmotically dependent steps by DNA uptake after Escherichia coli electroporation.

Currently, the major problem in the genetic transformation is to understand how such a large molecule as the plasmid DNA passes through the cell membrane. To solve this problem we used a simplified experimental model with Escherichia coli and the plasmid pBR322: the DNA-bacteria mixture was electroporated in a sucrose solution at 0 degree C and at fixed electrical parameters; the samples were then directly plated into agar. It was found that the electrically-induced bacterial transformation after pulsing is dependent on two factors: heat shock (delta T) and osmotic stress. Our results indicate that two mechanisms contribute to these effects: (i) thermodiffusion of the DNA across the membrane and (ii) osmotic flows of the medium, containing the DNA, into the interior of the cell.