Tan H, Yeung E S
Ames Laboratory-USDOE and Department of Chemistry, Iowa State University, 50011, USA.
Electrophoresis. 1997 Dec;18(15):2893-900. doi: 10.1002/elps.1150181529.
The influence of three classes of fluorescence labels including dipyrrometheneboron difluoride (BODIPY), energy transfer (ET) and conventional fluorescein and rhodamine (ABI), on DNA sequencing has been examined with laser-induced fluorescence detection and poly(ethylene oxide)-filled capillary electrophoresis. DNA sequencing fragments were generated by dye-labeled primer cycle-sequencing reactions in a hot-air thermal cycler. A parameter, relative-induced shift, was introduced to quantify the uniformity of electrophoretic mobilities of these fragments. BODIPY was found to have the smallest, but nonzero, effect for dye-induced nonuniformity. Although ET dyes provided the highest sensitivity due to their unique spectroscopic properties, they were found to lack photostability compared to BODIPY and ABI dyes. Characterization also brings out some important tips for selecting the suitable dye set for the two-channel ratio-based DNA base-calling method.
利用激光诱导荧光检测和填充聚环氧乙烷的毛细管电泳技术，研究了包括二吡咯亚甲基硼二氟化物（BODIPY）、能量转移（ET）以及传统的荧光素和罗丹明（ABI）在内的三类荧光标记物对DNA测序的影响。DNA测序片段通过在热空气热循环仪中进行的染料标记引物循环测序反应产生。引入了一个参数，即相对诱导位移，以量化这些片段电泳迁移率的均匀性。结果发现，BODIPY对染料诱导的不均匀性影响最小，但不为零。尽管ET染料因其独特的光谱特性提供了最高的灵敏度，但与BODIPY和ABI染料相比，它们缺乏光稳定性。表征还为基于双通道比率的DNA碱基识别方法选择合适的染料组提供了一些重要提示。
