DNA molecules can be transported through microchannels with help of electrophoresis and flow. Confinement of DNA molecules leads to elongation of their unconstrained equilibrium configuration when passing the microchannel. Application of electrical fields reduces the mobility and entails DNA trapping because of high gradients of the field due to a decrease in the channels’ magnitude. Microfluidic channels in polydimethylsiloxane (PDMS) were formed by soft replica molding technology combining micro- and nanofluidic features. The applicability of the hybrid micro- and nanofluidic PDMS structures for single molecule observation and manipulation was demonstrated by introducing single molecules of λ-DNA into the channels using optimized conditions for the applied potential and flow.