Improved quantitation of DNA curvature using ligation ladders

It is often desirable to estimate accurately the local shape of DNA molecul es. Such measurements are useful in understanding the intrinsic contributio n of DNA sequence to curvature, as well as in assessing the effects of chem ical modifications. We have been investigating the effects of asymmetric ph osphate neutralization on DNA shape using the well-characterized ligation l adder approach developed by Crothers and co-workers [D.M. Crothers and J. D rak (1992) Meth, Enzymol., 212, 46-71]. This technique is remarkably sensit ive to differences in DNA shape. We now report a general quantitative assay of DNA curvature that we have validated using a set of phased A, tract sta ndards. This approach allows simultaneous estimation of helix axis deflecti on magnitude and direction when a test sequence is monitored in at least th ree phasings relative to a reference A,, tract in short DNA duplexes. Analy sis using this improved approach confirms our published data on DNA curvatu re due to electrostatic effects.