Scherer J R, Kheterpal I, Radhakrishnan A, Ja W W, Mathies R A
Department of Chemistry, University of California, Berkeley 94720, USA.
Electrophoresis. 1999 Jun;20(7):1508-17. doi: 10.1002/(SICI)1522-2683(19990601)20:7<1508::AID-ELPS1508>3.0.CO;2-7.
We have constructed a rotary confocal fluorescence scanner and capillary array electrophoresis system that is designed to analyze over 1000 DNA sequencing or fragment sizing separations in parallel. Capillaries are arranged around the surface of a cylinder and a rotating objective in the middle of the cylinder excites and collects fluorescence from labeled DNA fragments as they pass the capillary detection window. The capillaries are pressure-filled with a replaceable matrix and the samples are electrokinetically injected in parallel from a stainless steel microtiter plate at the cathode end. We demonstrate that the instrument is capable of producing four-color data from all capillaries at a scan rate of 4 Hz (corresponding to a linear scan velocity of 121 cm/s). M13 sequencing data were obtained using a 128 capillary array mounted in half of the first quadrant of the scanner. In this initial run, read lengths greater than 500 bases were obtained in over 60% of the capillaries.
我们构建了一种旋转共聚焦荧光扫描仪和毛细管阵列电泳系统,该系统旨在并行分析1000多个DNA测序或片段大小分离。毛细管围绕圆柱体表面排列,圆柱体中间的旋转物镜在标记的DNA片段通过毛细管检测窗口时激发并收集荧光。毛细管中压力填充有可更换的基质,样品从阴极端的不锈钢微量滴定板中通过电动方式并行注入。我们证明该仪器能够以4 Hz的扫描速率(对应于121 cm/s的线性扫描速度)从所有毛细管中产生四色数据。M13测序数据是使用安装在扫描仪第一象限一半位置的128毛细管阵列获得的。在这次初始运行中,超过60%的毛细管获得了大于500个碱基的读长。