摘要
目的 探讨用脉冲场凝胶电泳(PFGE)分离大分子环状DNA。方法 使用脉冲场凝胶电泳仪器,设定不同电泳条件,以已知分子大小的环状DNA作为标准参照物,对不同环状DNA分子进行分离。结果 使用倒转电场凝胶电泳(FIGE)方式,用0.6％或0.5％琼脂糖凝胶,正向电压梯度9V/cm,脉冲时间2.0s,反向电压梯度9V/cm,脉冲时间1.0s,缓冲溶液温度14℃,电泳时间8h,适宜分离2～33kb环状DNA分子。结论 脉冲场凝胶电泳是分离大分子DNA的必须技术。然而,电泳时的条件设定,对是否能有效分离影响很大。为了有效分离大分子DNA,需要摸索实验,建立最适设定条件。
Objective To study the separation of closed circular DNA using pulsed - field electrophoresis. Methods With known molecular sizes of closed circular DNA as markers, different electrophoresis conditions to separate closed circular DNA were investigated. Results By FIGE mode, closed circular 2 kb - 33kb DNA were well separated with Forward 9 V/cm, Fst=2.0s, Reverse 6 V/cm, Rst= 1.0 s, 0.5× TBE Buffer, Buffer temperature 14℃, Run time 8 hours. Conclusion Pulsed - field electrophoresis is an essential technique to separate large DNA. The electrophoretic condition exerts a considerable influence on experiment results. *
出处
《临床和实验医学杂志》
2003年第3期138-141,共4页
Journal of Clinical and Experimental Medicine
关键词
分离
环状DNA分子
脉冲场凝胶电泳
倒转电场凝胶电泳
脉冲时间
Pulsed-field
Pulse time
FIGE (Field Inversion Gel Electrophoresis)
CHEF (Contour-clamped Homoge-neous Electric Field Gel Electrophoresis)
Closed circular DNA
