Pulsed-field electrophoresis of nucleic acids: ultrafast separations in ultrashort capillaries

Michael D. Morris; Yongseong Kim; Richard W. Hammond

doi:10.1117/12.237609

1 April 1996 Pulsed-field electrophoresis of nucleic acids: ultrafast separations in ultrashort capillaries

Michael D. Morris, Yongseong Kim, Richard W. Hammond

Proceedings Volume 2680, Ultrasensitive Biochemical Diagnostics; (1996) https://doi.org/10.1117/12.237609
Event: Photonics West '96, 1996, San Jose, CA, United States

Abstract

Nucleic acid electrophoretic separations in unentangled water-soluble polymers are rapid and applicable over the length range 500 bp to greater than 1.5 Mbp, depending upon the polymer matrix chosen. We have been able to achieve high resolution separations in 3 - 4 minutes, even in the Mbp size range. Pulsed field electrophoresis is necessary for chains longer than about 20 kbp and is beneficial above about 2 - 3 kbp. The separation speed is a consequence of several factors: (1) the low viscosity of an unentangled polymer solution. (2) the high electric fields usable in pulsed field electrophoresis. And (3) the use of short (ca. 10 cm) capillaries. Electrophoretic measurements are guided by video fluorescence microscopy, which is used to study the dynamics of the separation process. Analysis of image sequences demonstrates that the segmental motions of DNA, which generate the size-dependent mobilities, are quite similar to those previously observed in entangled linear polymers and cross-linked gels. We describe our most recent separations and the current state of understanding of DNA dynamics in unentangled polymer matrices.

Citation Download Citation

Michael D. Morris, Yongseong Kim, and Richard W. Hammond "Pulsed-field electrophoresis of nucleic acids: ultrafast separations in ultrashort capillaries", Proc. SPIE 2680, Ultrasensitive Biochemical Diagnostics, (1 April 1996); https://doi.org/10.1117/12.237609

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