使用具有多波长荧光检测功能的毛细管电泳对两个多重短串联重复序列系统进行分析。

Analysis of two multiplexed short tandem repeat systems using capillary electrophoresis with multiwavelength fluorescence detection.

作者信息

Isenberg A R, Allen R O, Keys K M, Smerick J B, Budowle B, McCord B R

机构信息

Department of Chemistry, University of Virginia, Charlottesville, USA.

出版信息

Electrophoresis. 1998 Jan;19(1):94-100. doi: 10.1002/elps.1150190117.

DOI:10.1002/elps.1150190117

PMID:9511869

Abstract

A series of experiments was performed to analyze the utility of capillary electrophoresis (CE) with multiwavelength detection capabilities for multiplex typing of short tandem repeat loci. Characteristics of the sieving polymer, hydroxyethylcellulose, which affect resolution of single strand (ss) DNA fragments were examined. Additionally, the effects of denaturant in the polymer system, separation voltage, and analysis temperature were studied to ascertain their effects on DNA separations and capillary lifetime. The use of elevated run temperature (60 degrees C) was found to improve sizing precision, to increase the lifetime of capillaries (100 runs or more per capillary), and to provide runtimes of under 20 min. Finally, 100 individual human DNA samples were typed successfully using CE. The average resolution obtained was 1.4 bases for a 200 base fragment with a standard deviation of sizing of 0.2 bases, allowing all alleles examined to be distinguished clearly.

摘要

进行了一系列实验，以分析具有多波长检测能力的毛细管电泳（CE）用于短串联重复序列位点多重分型的效用。研究了影响单链（ss）DNA片段分辨率的筛分聚合物羟乙基纤维素的特性。此外，还研究了聚合物体系中的变性剂、分离电压和分析温度对DNA分离和毛细管寿命的影响。发现使用升高的运行温度（60摄氏度）可提高大小测定精度，增加毛细管寿命（每根毛细管运行100次或更多次），并使运行时间在20分钟以内。最后，使用CE成功地对100个人类个体DNA样本进行了分型。对于200个碱基的片段，获得的平均分辨率为1.4个碱基，大小测定的标准偏差为0.2个碱基，使得所有检测的等位基因都能被清晰区分。