Legerski R J, Gray H B
Biochim Biophys Acta. 1976 Aug 18;442(2):129-41. doi: 10.1016/0005-2787(76)90483-4.
A method is described for the rapid separation of the complementary strands of homogeneous DNA's. The method takes advantage of a difference in sedimentation coefficients between the complementary strands when these have been dissociated and complexed with poly(U-G). The separation procedure is readily carried out for preparative purposes by zone sedimentation in linear 5--20% (w/v) sucrose gradients containing 2 M NaCl. In analytical band sedimentation experiments, concentrated solutions of CsCl have been used to effect the separation of the complexed complementary strands. Sedimentation coefficients corresponding to corrected values at infinite dilution for the cesium salt of the nucleic acid complexes at 25 degrees C (s 25, w) have been obtained for the complexed complementary strands of coliphage lambdab2b5c DNA and for the uncomplexed single strands of this DNA. The relative values of the parameters affecting s 25, w have been determined. Under these conditions, for which preferential hydration of the sedimenting species is not involved, it was found that the more highly complexed strand has sustained a 24% increase in mass and a 28% decrease in frictional coefficient relative to uncomplexed DNA, while the strand binding the lesser amount of RNA has increased in mass by 11% and changed in frictional coefficient by an insignificant amount relative to the uncomplexed species from that of the uncomplexed DNA do not contribute significantly to the differences in s 25, w between the complexed complementary strands.
本文描述了一种快速分离均一DNA互补链的方法。该方法利用了互补链与聚（U-G）解离并复合后沉降系数的差异。通过在含2M氯化钠的线性5%-20%（w/v）蔗糖梯度中进行区带沉降，可方便地进行制备性分离操作。在分析性区带沉降实验中，已使用浓氯化铯溶液来实现复合互补链的分离。已获得了25℃下噬菌体λb2b5c DNA复合互补链及该DNA未复合单链核酸复合物铯盐在无限稀释时校正值对应的沉降系数（s25,w）。已确定了影响s25,w的参数的相对值。在不涉及沉降物质优先水合作用的这些条件下，发现与未复合DNA相比，复合程度更高的链质量增加了24%，摩擦系数降低了28%，而结合较少RNA的链质量增加了11%，摩擦系数相对于未复合链变化不显著。未复合DNA的那些参数对复合互补链之间s25,w的差异贡献不大。
