Microvolume quantitation of nucleic acids.
Microvolume quantitation of nucleic acids.
复制标题
DOI：
10.1002/0471142727.mba03js93
复制
发表时间：
2011-01-01
影响因子：
--
通讯作者：
Conklin, Deborah S
中科院分区：
文献类型：
--
作者：
Desjardins, Philippe R;Conklin, Deborah S
几十年来，通过吸光度和荧光光谱法定量DNA和RNA一直是生命科学中的有力工具。核酸定量的经典方法需要用样品填充装置，例如比色皿和毛细管（传统方法描述见附录3D）。随着越来越多的分子生物学技术产生越来越少量的分离样品，并且需要以最小的样品消耗精确定量核酸，微体积样品的分析已经变得至关重要。光子技术的进步催生了一种开创性的微体积系统，该系统将光纤技术与样品的固有物理特性相结合，可大幅减少测量体积，从而消除了对比色皿和毛细管的需求。自从第一台微体积仪器问世以来，现在有几种新的设计，提供了使用更少量的材料测量核酸的机会。总之，这些系统不仅减少了测量体积（小至0.5至2升），而且往往比传统方法更有效，即使在样品丰富的情况下也很有用。本单元中的方案基于最广泛接受的微体积系统，并旨在作为传统核酸定量方法的实际替代方案。
Quantitation of DNA and RNA by absorbance and fluorescence spectroscopy has been a powerful tool in life sciences for decades. Classic methods of nucleic acid quantitation require the filling of devices, such as cuvettes and capillaries, with sample (traditional methodologies are described in APPENDIX 3D). Analysis of microvolume samples has become of paramount importance as more molecular biology techniques yield progressively smaller amounts of isolated sample and require accurate quantitation of nucleic acids with minimal consumption of sample. Advances in photonic technologies have resulted in a pioneering microvolume system that combines fiber optic technology with the inherent physical properties of the sample to dramatically reduce measurement volumes, removing the need for cuvettes and capillaries. Since the introduction of the first microvolume instrument, several new designs are now available, providing opportunities to measure nucleic acids using much smaller amounts of material. Altogether, these systems not only reduce measurement volume (as little as 0.5 to 2 l), but also tend to be more efficient time-wise than traditional methods, making them useful even when sample is plentiful. The protocols in this unit are based on the most widely accepted microvolume systems and are intended as practical alternatives to traditional nucleic acid quantitation methodology.