Three polyether-tethered berberine dimers (1a–c) were studied for their binding affinity, selectivity and thermal stabilization towards human telomeric dimeric quadruplex DNA (G2T1). Compound 1a with the shortest polyether linker showed the highest affinity (K_a > 10⁸ M⁻¹) and 76–508-fold higher selectivity for mixed-type G2T1 over antiparallel G2T1 and three monomeric G-quadruplexes, which are human telomeric monomeric quadruplex G1, c-kit 1 and c-kit 2. Compound 1a induced the formation of quadruplex structures and showed higher thermal stabilization for mixed-type G2T1 than for anti-parallel G2T1, G1 and ds DNA. Spectroscopic studies suggest that compound 1a could bind to mixed-type G2T1 via end-stacking and external binding modes. These results suggest that the polyether linkers in these compounds play an important role in regulating the binding affinity and selectivity towards mixed-type G2T1 and that compound 1a could target mixed-type G2T1 at other genome regions with antiparallel G2T1 and monomeric G-quadruplexes. These results may provide useful guidance for the rational design of selective multimeric G-quadruplex binders and potential anticancer agents.