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Published August 11, 1988 | Published
Journal Article Open

Optimized conditions for pulsed field gel electrophoretic separations of DNA


Quantitative measurement of DNA migration in gel electrophoresis requires precisely controlled homogeneous electric fields. A new electrophoresis system has allowed us to explore several parameters governing DNA migration during homogeneous field pulsed field gel (PFG) electrophoresis. Migration was measured at different switch times, temperatures, agarose concentrations, and voltage gradients. Conditions which increase DNA velocities permit separation over a wider size range, but reduce resolution. We have also varied the angle between the alternating electric fields. Reorientation angles between 105° and 165° give equivalent resolution, despite significant differences in DNA velocity. Separation of DNA fragments from 50 to greater than 7000 kilobases (Kb) can easily be optimized for speed and resolution based on conditions we describe.

Additional Information

© 1988 IRL Press, Limited. Received February 29, 1988; Revised and Accepted June 28, 1988. We would like to thank Doug Vollrath for providing a copy of his manuscript prior to publication, John Carbon for providing s. pombe probes, Charles Spence for software development, Petros Arakelian for technical assistance, and Hiroaki Shizuya for his critical reading of the manuscript. This work was supported by grants from the NSF to L.H. (DMB 85-00298) and from NIH to M.I.S. (GM 34236). B.W.B. is a NIH postdoctoral fellow (GM 10974).

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