The interaction of calf thymus DNA(ctDNA) with berberinc(Ber) was systematically investigated via UV-Vis,fluorescence and phosphorescence spectroscopies in aqueous solution and on solid substrate.The very weak fluorescence intensity of Ber at 525 nm in aqueous solution can be greatly enhanced by ctDNA.The UV-Vis spectrum shows that Ber could bind with DNA by intercalation.In addition,strong fluorescence of Ber alone could be observed on filter paper substrate and can be quenched by ctDNA effectively,and the increase of fluorescence polarization of Ber on the filter paper implies the intercalation binding.Ber emits phosphorescence at 619 nm in the presence of thallium(Ⅰ) acetate.The lifetime of Ber increased from 38.4 ms to 43.4 ms with the increase of ctDNA.The increase of lifetime is possibly attributed to the increase of rigidity of Ber after its intercalating into the ctDNA base pairs.This explanation is also augmented by the anionic quenching results.Ber behaves as a typical intercalating agent into ctDNA.The intrinsic binding constant between Ber and ctDNA is (1.84±0.12)× 104 L/mol and n=0.16.