The acyclic bis-phenanthridinium ligands 1, 2 and the cyclic analogue 3 bind to ss-RNA by bis-intercalation. Due to it's shorter linker 1 exhibits mono-intercalative binding to ds-polynucleotides, while a mixed mode of binding with 2 is shown to strongly dependent on the base composition and tertiary structure of ds-DNA and RNA. The cyclic analogue 3 binds to ds-polynucleotides by non-intercalative mode. Comparing the ss-/ds-polynucleotide selectivity obtained for 3 and previously reported for 3,8-linked bis-phenanthridinium analogues, it is clear that the more rigid structure and sterically more restricted cleft of the latter could better distinguish ss- from ds-polynucleotide regions.