二价锰离子导致的DNA构象转变实验研究

发布时间：2018-03-25 18:27

本文选题：DNA构象　切入点：Mn~(2+)　出处：《温州大学》2016年硕士论文

【摘要】：在本文中,我们利用单分子磁镊(Magnetic Tweezers,MT)和原子力显微镜(Atomic Force Microscope,AFM)直观的看到了二价锰离子导致DNA凝聚的现象。单分子磁镊实验可以排除DNA分子间的相互作用直观的表明在室温条件下Mn~(~(2+))可以导致DNA凝聚。为了研究是否所有的实验体系均有这种实验现象,我们使用原子力显微镜观察了在相同缓冲液条件下Mn~(~(2+))导致的l-DNA的形貌构象。实验结果表明,当锰离子浓度为1mM时,DNA的形貌为典型的线团状结构,当锰离子的浓度增加为3mM时,DNA的形貌为典型的凝聚状,表明DNA发生了凝聚。曼宁理论指出二价金属离子仅能中和DNA 88%的负电荷,而DNA凝聚现象需要中和掉90%以上电荷才可发生,这种现象是曼宁理论不能解释的。这是因为曼宁理论是以扩散结合的为前提的理论。曼宁理论的扩散结合指出结合的抗衡离子可以在给定的区域自由流动。碱土金属离子与DNA的结合是通过离子与DNA链磷酸之间的静电吸引,这符合扩散结合的特点。而已有的研究表明,Mn~(~(2+))更偏向于与DNA的G碱基上的N7原子结合甚至可以改变DNA的结构。这表明,Mn~(~(2+))与DNA相互作用时局部有强烈的静电模式,因此自由流动的Mn~(~(2+))是有限的不符合扩散结合的特点,因此曼宁理论无法解释目前的实验结果。为了研究温度对DNA凝聚的影响,我们使用单分子磁镊方法研究了55°C条件下对DNA凝聚的影响,实验表明升高温度对单分子DNA凝聚无明显影响;同样我们用原子力显微镜观察了温度对DNA形貌的影响,实验表明随着温度的升高有助于DNA的凝聚但并不是使DNA凝聚的必要条件。
[Abstract]:In this article, Using the single molecular magnetic tweezers (MMT) and atomic force microscope (AFM), we can directly see the phenomenon of DNA condensation induced by manganese ions. The experiment of single molecular magnetic tweezers can eliminate the interaction between DNA molecules and show that at room temperature the interaction between DNA molecules can be ruled out intuitively by the experiment of monolayer magnetic tweezers and atomic force microscope (AFM). In order to study whether all experimental systems have this kind of experimental phenomenon, The morphologic conformation of l-DNA induced by Mn~(~(2 in the same buffer solution was observed by atomic force microscope. The experimental results show that the morphology of l-DNA is a typical coiled structure when the concentration of mn ~ (2 +) is 1mM. When the concentration of manganese ion is increased to 3mM, the morphology of DNA is typical, which indicates that the condensation occurs. Manning theory indicates that bivalent metal ions can neutralize only 88% negative charges of DNA, while DNA condensation needs to neutralize more than 90% of the charge. This phenomenon can not be explained by Manning theory. This is because the Manning theory is based on diffusion binding theory. The diffusion combination of Manning theory indicates that the counterweight ions can flow freely in a given region. Metal ions bind to DNA through electrostatic attraction between ions and DNA chain phosphoric acid. This is in line with the characteristics of diffusion binding. However, some studies have shown that the N7 atom on the G base of DNA may even change the structure of DNA, which indicates that there is a strong local electrostatic mode in the interaction between MN and DNA. Therefore, the free-flowing Mn~(~(2) is a finite nonconforming to the characteristics of diffusion bonding. Therefore, Manning's theory cannot explain the present experimental results. In order to study the effect of temperature on DNA condensation, The effect of temperature on DNA condensation at 55 掳C was studied by using single molecular magnetic tweezers. The experimental results show that the increase of temperature has no obvious effect on the DNA condensation of single molecule, and the effect of temperature on the morphology of DNA is also observed by atomic force microscope (AFM). The experimental results show that with the increase of temperature, the condensation of DNA is helpful, but it is not a necessary condition for DNA to agglomerate.
【学位授予单位】：温州大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：O629.74

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