Scanning force microscopy of DNA deposited onto mica: Equilibration versus kinetic trapping studied by statistical polymer chain analysis
Scanning force microscopy of DNA deposited onto mica: Equilibration versus kinetic trapping studied by statistical polymer chain analysis
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DOI：
10.1006/jmbi.1996.0687
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发表时间：
1996-12-20
影响因子：
5.6
通讯作者：
Bustamante, C
中科院分区：
文献类型：
--
作者：
Rivetti, C;Guthold, M;Bustamante, C
本文报道了在扫描力显微镜（SFM）下研究DNA分子在云母表面沉积成像的过程。动力学实验表明，DNA分子从溶液滴到表面的运输完全由扩散控制，并且分子被不可逆地吸附到底物上。统计聚合物链分析已应用于DNA分子，以确定导致平衡的沉积条件和导致被困构型的沉积条件。在适当的条件下，DNA分子沉积在新切割的云母上，能够像在理想的二维溶液中一样在表面上平衡。这些分子的滞留长度为53 nm。尽管有额外的蛋白质-表面相互作用，但在两端标记有辣根过氧化物酶链亲和素融合蛋白的DNA片段仍然能够在表面上保持平衡。相比之下，DNA分子沉积在发光云母或H+交换云母表面不平衡。这些分子的构象类似于表面上的简单投影，这表明了一个动力学捕获过程。这些结果验证了最近SFM用于定量分析沉积在云母上的复杂大分子组合的构象的应用。在平衡条件下，本研究表明，SFM可以用于确定DNA分子的持续长度，精度很高。(C) 1996学术出版社有限公司
This paper reports a study of the deposition process of DNA molecules onto a mica surface for imaging under the scanning force microscope (SFM). Kinetic experiments indicate that the transport of DNA molecules from the solution drop onto the surface is governed solely by diffusion, and that the molecules are irreversibly adsorbed onto the substrate. A statistical polymer chain analysis has been applied to DNA molecules to determine the deposition conditions that lead to equilibrium and those that result in trapped configurations. Using the appropriate conditions, DNA molecules deposited onto freshly cleaved mica, are able to equilibrate on the surface as in an ideal two-dimensional solution. A persistence length of 53 nm was determined from those molecules. DNA fragments that were labeled on both ends with a horseradish peroxidase streptavidin fusion protein were still able to equilibrate on the surface, despite the additional protein-surface interaction. In contrast, DNA molecules deposited onto glow-discharged mica or H+ -exchanged mica do not equilibrate on the surface. These molecules adopt conformations similar to those expected for a simple projection onto the surface plane, suggesting a process of kinetic trapping.These results validate recent SFM application to quantitatively analyze the conformation of complex macromolecular assemblies deposited on mica. Under equilibration conditions, the present study indicates that the SFM can be used to determine the persistence length of DNA molecules to a high degree of precision. (C) 1996 Academic Press Limited.