Abstract
Pulsed-field capillary electrophoresis represents a new tool for rapid and highly efficient separations of large biopolymers. The method has been utilized here to study dependencies of the electrophoretic mobility upon the frequency and pulse shape of applied voltage for large, double-stranded DNA molecules (5-100 kb) migrating in neutral polymer solutions. Two different shapes of alternating electric field (sine- and square-wave impulses) were examined with the frequency values ranging from 1 to 30 Hz. The linear dependence between duration of the forward pulse (at which the DNA molecule experiences a minimum mobility) and the product N.In(N) (where N is the number of base pairs) was experienced in field-inversion gel electrophoresis, while exponential dependence was found with the sinusoidal electric field. The mobility minima were lower in field-inversion electrophoresis than with the biased sinusoidal-field technique. The DNA (5 kb concatamers) was adequately separated using a ramp of frequency in the square-wave electric field, in approximately 1 h. The migration order of DNA fragments was referenced through adding a monodisperse DNA (48.5 kb) into the sample. The band inversion phenomena were not observed under any experimental conditions used in this work.
Full text
PDF





Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Carle G. F., Frank M., Olson M. V. Electrophoretic separations of large DNA molecules by periodic inversion of the electric field. Science. 1986 Apr 4;232(4746):65–68. doi: 10.1126/science.3952500. [DOI] [PubMed] [Google Scholar]
- Demana T., Lanan M., Morris M. D. Improved separation of nucleic acids with analyte velocity modulation capillary electrophoresis. Anal Chem. 1991 Dec 1;63(23):2795–2797. doi: 10.1021/ac00023a023. [DOI] [PubMed] [Google Scholar]
- Duke TA. Tube model of field-inversion electrophoresis. Phys Rev Lett. 1989 Jun 12;62(24):2877–2880. doi: 10.1103/PhysRevLett.62.2877. [DOI] [PubMed] [Google Scholar]
- Guszczynski T., Pulyaeva H., Tietz D., Garner M. M., Chrambach A. Capillary zone electrophoresis of large DNA. Electrophoresis. 1993 May-Jun;14(5-6):523–530. doi: 10.1002/elps.1150140180. [DOI] [PubMed] [Google Scholar]
- Heiger D. N., Cohen A. S., Karger B. L. Separation of DNA restriction fragments by high performance capillary electrophoresis with low and zero crosslinked polyacrylamide using continuous and pulsed electric fields. J Chromatogr. 1990 Sep 7;516(1):33–48. doi: 10.1016/s0021-9673(01)90202-x. [DOI] [PubMed] [Google Scholar]
- Kotaka T., Adachi S., Shikata T. Biased sinusoidal field gel electrophoresis for the separation of large DNA. Electrophoresis. 1993 Apr;14(4):313–321. doi: 10.1002/elps.1150140154. [DOI] [PubMed] [Google Scholar]
- Liu J. P., Hsieh Y. Z., Wiesler D., Novotny M. Design of 3-(4-carboxybenzoyl)-2-quinolinecarboxaldehyde as a reagent for ultrasensitive determination of primary amines by capillary electrophoresis using laser fluorescence detection. Anal Chem. 1991 Mar 1;63(5):408–412. doi: 10.1021/ac00005a004. [DOI] [PubMed] [Google Scholar]
- McDonell M. W., Simon M. N., Studier F. W. Analysis of restriction fragments of T7 DNA and determination of molecular weights by electrophoresis in neutral and alkaline gels. J Mol Biol. 1977 Feb 15;110(1):119–146. doi: 10.1016/s0022-2836(77)80102-2. [DOI] [PubMed] [Google Scholar]
- Noolandi J, Rousseau J, Slater GW, Turmel C, Lalande M. Self-trapping and anomalous dispersion of DNA in electrophoresis. Phys Rev Lett. 1987 Jun 8;58(23):2428–2431. doi: 10.1103/PhysRevLett.58.2428. [DOI] [PubMed] [Google Scholar]
- Novotny M. V., Cobb K. A., Liu J. P. Recent advances in capillary electrophoresis of proteins, peptides and amino acids. Electrophoresis. 1990 Sep;11(9):735–749. doi: 10.1002/elps.1150110911. [DOI] [PubMed] [Google Scholar]
- Orbach M. J., Vollrath D., Davis R. W., Yanofsky C. An electrophoretic karyotype of Neurospora crassa. Mol Cell Biol. 1988 Apr;8(4):1469–1473. doi: 10.1128/mcb.8.4.1469. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schwartz D. C., Cantor C. R. Separation of yeast chromosome-sized DNAs by pulsed field gradient gel electrophoresis. Cell. 1984 May;37(1):67–75. doi: 10.1016/0092-8674(84)90301-5. [DOI] [PubMed] [Google Scholar]
- Stefansson M., Novotny M. Separation of complex oligosaccharide mixtures by capillary electrophoresis in the open-tubular format. Anal Chem. 1994 Apr 1;66(7):1134–1140. doi: 10.1021/ac00079a031. [DOI] [PubMed] [Google Scholar]
- Sudor J., Novotny M. V. Pulsed-field capillary electrophoresis: optimizing separation parameters with model mixtures of sulfonated polystyrenes. Anal Chem. 1994 Jul 1;66(13):2139–2147. doi: 10.1021/ac00085a032. [DOI] [PubMed] [Google Scholar]
- Sudor J., Novotny M. V. Separation of large DNA fragments by capillary electrophoresis under pulsed-field conditions. Anal Chem. 1994 Aug 1;66(15):2446–2450. doi: 10.1021/ac00087a007. [DOI] [PubMed] [Google Scholar]
- Tokunaga M., Wada N., Hishinuma F. Expression and identification of immunity determinants on linear DNA killer plasmids pGKL1 and pGKL2 in Kluyveromyces lactis. Nucleic Acids Res. 1987 Feb 11;15(3):1031–1046. doi: 10.1093/nar/15.3.1031. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Viovy J. L., Duke T. DNA electrophoresis in polymer solutions: Ogston sieving, reptation and constraint release. Electrophoresis. 1993 Apr;14(4):322–329. doi: 10.1002/elps.1150140155. [DOI] [PubMed] [Google Scholar]
