Reed J, Singer E, Kresbach G, Schwartz D C
Department of Chemistry, W. M. Keck Laboratory for Biomolecular Imaging, New York University, New York 10003, USA.
Anal Biochem. 1998 May 15;259(1):80-8. doi: 10.1006/abio.1998.2640.
Many new techniques in biomolecular chemistry and genomic analysis require the immobilization of molecular reagents on specially prepared surfaces. However, the process of molecular fixation often interferes with or precludes the use of standard in vitro biochemical assays. Optical mapping is an emergent technology for genomic analysis which relies on the biochemical activity of DNA fixed to silanized glass surfaces. Optical mapping surfaces have been shown to be compatible with restriction endonucleases and a variety of DNA polymerases. The essential properties of biochemically active surfaces are poorly understood in most of the current technologies which utilize molecular fixation, including optical mapping. The purpose of this study is to use the powerful technique of atomic force microscopy, in combination with informative enzymatic assays, to correlate biochemical activity with microscopic surface structure. The results presented provide meaningful insight into the effect of surface preparation on the biochemical accessibility of surface-bound molecules. Novel analysis which may facilitate the automation of optical mapping is presented.
生物分子化学和基因组分析中的许多新技术都需要将分子试剂固定在特制的表面上。然而，分子固定过程常常会干扰或排除标准体外生化分析的使用。光学图谱分析是一种新兴的基因组分析技术，它依赖于固定在硅烷化玻璃表面的DNA的生化活性。光学图谱分析表面已被证明与限制性内切酶和多种DNA聚合酶兼容。在目前大多数利用分子固定的技术中，包括光学图谱分析，对具有生化活性表面的基本特性了解甚少。本研究的目的是结合强大的原子力显微镜技术和信息丰富的酶分析方法，将生化活性与微观表面结构联系起来。所呈现的结果为表面制备对表面结合分子生化可及性的影响提供了有意义的见解。本文还提出了可能有助于光学图谱分析自动化的新颖分析方法。
