The hybridization performance of a set of 12-mer RNA:RNA duplexes containing 2'-C-methyluridine, 5-bromo-2'-C-methyluridine, or (2'S)-2'-deoxy-2'-C-methyluridine was analyzed. Melting point temperatures of the modified duplexes showed an important ΔT(m) decrease (-8.9 to -12.5 °C), while circular dichroism experiments indicated that the helix was still A-type, suggesting a localized disturbance disorder. Molecular dynamics simulations using AMBER were carried out in order to gain structural knowledge about the effect of the 2'-C-methyl modification in double stranded environments. On the other hand, in an attempt to explain the behavior of the 2'-deoxy-2'-C-methyl nucleosides in single stranded environments, like the 10-23 DNAzyme core, molecular dynamic simulations were performed, incorporating the modified analogues into single stranded reported stem-loop structures, studding the sugar conformations along the MD trajectories. It was observed that, despite their preferential conformational states, the 2'-C-methyl analogues are flexible enough to adopt a different puckering in single stranded environments.