Anderson A C, Scaringe S A, Earp B E, Frederick C A
Committee on Biophysics at Harvard University, and Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA.
RNA. 1996 Feb;2(2):110-7.
Homogeneous preparations of milligram quantities of RNA are a prerequisite for their characterization by biophysical methods such as crystallography or NMR spectroscopy. Methods for obtaining milligram quantities of pure synthetic RNA are described in this paper. These methods employ anion exchange HPLC for purifying full-length sequence from failure sequences and incompletely deprotected material. RNA molecules with little or extensive amounts of secondary structure could be purified. In cases where the RNA molecule was tightly folded, the cation in the eluent buffer influenced both the distinction of the peaks during chromatography and the final folded conformation. Finally, two RNA sequences were chemically synthesized, deprotected, purified, and crystallized using this methodology.
毫克量RNA的均一制剂是通过诸如晶体学或核磁共振光谱等生物物理方法对其进行表征的前提条件。本文描述了获得毫克量纯合成RNA的方法。这些方法采用阴离子交换高效液相色谱从失败序列和未完全脱保护的材料中纯化全长序列。几乎没有二级结构或具有大量二级结构的RNA分子都可以被纯化。在RNA分子紧密折叠的情况下，洗脱缓冲液中的阳离子会影响色谱过程中峰的区分以及最终的折叠构象。最后，使用该方法化学合成、脱保护、纯化并结晶了两个RNA序列。
