單分子光鉗系統之建置校正及生醫應用

Setup of Single-Molecule Optical Tweezers and Its Applications in Biomedical Research

單分子光鉗技術可施加pN大小的力於生物系統中，隨著解析度不斷提升，甚至已經能準確地偵測到原子大小等級的距離變化，其技術廣泛的用於生物體系下分子馬達，DNA、RNA及蛋白質折疊，DNA-蛋白質作用力的研究。本篇描述單分子光鉗之架設與校正方法，同時介紹兩種常用的生物物理實驗方法：（1）拉伸實驗搭配聚合物模型分析，以及討論用於描述聚合物（DNA、RNA、蛋白質）模型的適用範圍；（2）被動式定力鉗則可用於即時的結構變化量測，不需要額外回饋來維持定力，有效提高了實驗量測上的時間與空間解析度。
Single-molecule optical tweezers platform applies pico-Newton size of forces and has been widely used to study the functional mechanisms in biological systems, such as the motion of molecular motors, folding of DNA or RNA and protein-DNA interactions in the spatial resolution of few A. Here, we described the detailed guide for instrumental building and calibrations of high-resolution optical tweezers and two types of commonly-used experimental assays. First, the force-extension assay can measure the extension under applied force by stretching and relaxing the molecule, and analyze the contour length and persistence length. Second, the passive force-clamp operates in a nearly zero trap-stiffness region and therefore, removes the needs to actively feedback the system to maintain a constant force throughout the experiments. Hence, the passive force-clamp platform can be used to study fast dynamics with small length change.