Yamaguchi Yoshinori, Todorov Todor I, Morris Michael D, Larson Ronald G
Department of Chemistry, University of Michigan, Ann Arbor, MI 48109-1055, USA.
Electrophoresis. 2004 Apr;25(7-8):999-1006. doi: 10.1002/elps.200305783.
The distribution of center of mass electrophoretic mobility mobilities and normalized migration time of up to 1080 lambda DNA molecules per experiment were measured in both semidilute hydroxyethylcellulose HEC/0.5 x Tris-borate-EDTA (TBE) solutions and dilute HEC/0.5 x TBE solution by high-speed video microscopy. Measurements were made microscopically over a short migration distance in homogeneous DNA HEC/0.5 x TBE solution and after electrophoretic migration of a plug of DNA through 7 cm. Video at 120 frames/s (semidilute HEC solution) and 236 frames/s (dilute HEC solution) allowed visualization with adequate resolution for single molecule mobility measurements. The electrophoretic migration times and band shapes predicted from the measurements corresponded well with those measured by conventional capillary electrophoresis (CE) in both semidilute and dilute HEC. In semidilute solution, the band width predicted by a square root of time scaling is in good agreement with the results of conventional CE. However, in dilute solution the precision of the measurements was not good enough to allow scaled estimates of band widths.
通过高速视频显微镜,在半稀羟乙基纤维素(HEC)/0.5×Tris-硼酸盐-EDTA(TBE)溶液和稀HEC/0.5×TBE溶液中,测量了每次实验多达1080个λDNA分子的质心电泳迁移率和归一化迁移时间的分布。测量是在均匀的DNA HEC/0.5×TBE溶液中短迁移距离上以及DNA凝胶块电泳迁移7厘米后通过显微镜进行的。120帧/秒(半稀HEC溶液)和236帧/秒(稀HEC溶液)的视频能够以足够的分辨率进行可视化,以测量单分子迁移率。在半稀和稀HEC中,测量预测的电泳迁移时间和条带形状与传统毛细管电泳(CE)测量的结果非常吻合。在半稀溶液中,由时间平方根标度预测的条带宽度与传统CE的结果吻合良好。然而,在稀溶液中,测量精度不够高,无法对标度后的条带宽度进行估计。