BUOYANT DENSITY OF DNA HOECHST-33258 (BISBENZIMIDE) COMPLEXES IN CSCL GRADIENTS - HOECHST-33258 BINDS TO SINGLE AT BASE-PAIRS

Buoyant density of DNA in CsCl gradients with Hoechst 33258 (bisbenzimide) was investigated as a function of guanine plus cytosine content of the DNA (%GC; in mole percent). A formula for calculating %GC from the refractive index (nd) of the isopycnic CsCl/Hoechst 33258 solution over the range of 0-75 %GC was established: %GC = 351762.28 × nd -123778.66 × nd 2 -249789.47 (the coefficients must not be rounded off). The shape of this curve indicates that under these conditions, in contrast to dilute buffers, Hoechst 33258 binds to single AT base pairs on DNA. Resolution of DNA bands in CsCl/Hoechst 33258 gradients is 1.6 to 2.1 times better than comparative CsCl gradients without the dye. Potential application to %GC determination is discussed. © 1991.