Nielsen P E, Zhen W P, Henriksen U, Buchardt O
Department of Biochemistry B, Panum Institute, Copenhagen, Denmark.
Biochemistry. 1988 Jan 12;27(1):67-73. doi: 10.1021/bi00401a012.
We have found that di-, tri-, tetra-, and hexa-9-acridinylamines are so efficiently associated with DNA during electrophoresis in polyacrylamide or agarose gels that they retard its migration. The retardation is roughly proportional to the reagent to base pair ratio, and the magnitude of the retardation indicates that a combined charge neutralization/helix extension mechanism is mainly responsible for the effect. Furthermore, DNA sequence dependent differences are observed. Thus, the pUC 19 restriction fragments (HaeIII or AluI), which in the native state comigrate upon gel electrophoretic analysis, could be separated in the presence of a diacridine, and specific DNA fragments responded differently to different diacridines. These results suggest that the effect also is due to a contribution from the DNA conformation and that the DNA conformation dynamics are influenced differently upon binding of different diacridines. We foresee three applications of this observation: (1) in analytical gel electrophoretic separation of otherwise comigrating DNA molecules, (2) in studies of polyintercalator-DNA interaction, and (3) in measurements of polyintercalator-induced DNA unwinding.
我们发现,二 - 、三 - 、四 - 和六 - 9 - 吖啶基胺在聚丙烯酰胺或琼脂糖凝胶电泳过程中与DNA的结合效率极高,以至于会阻碍其迁移。这种阻碍大致与试剂与碱基对的比例成正比,并且阻碍的程度表明电荷中和/螺旋延伸的联合机制是造成这种效应的主要原因。此外,还观察到了DNA序列依赖性差异。因此,在凝胶电泳分析中天然状态下共迁移的pUC 19限制性片段(HaeIII或AluI),在二吖啶存在的情况下可以被分离,并且特定的DNA片段对不同的二吖啶有不同的反应。这些结果表明,这种效应也归因于DNA构象的作用,并且不同的二吖啶结合后对DNA构象动力学的影响不同。我们预见到这一观察结果有三个应用:(1)用于分析凝胶电泳分离原本共迁移的DNA分子,(2)用于研究多嵌入剂与DNA的相互作用,(3)用于测量多嵌入剂诱导的DNA解旋。