Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1989 Sep 25;17(18):7359–7369. doi: 10.1093/nar/17.18.7359

Pulse time and agarose concentration affect the electrophoretic mobility of cccDNA during PFGE and FIGE [corrected].

B W Sobral 1, A G Atherly 1
PMCID: PMC334815  PMID: 2798097

Abstract

Circular DNAs have been shown to migrate in an unusual manner during field inversion gel electrophoresis (FIGE) and orthogonal field alternating gel electrophoresis (OFAGE). We studied the effect of varying pulse time and agarose concentration on the electrophoretic mobility of supercoiled (ccc) DNAs ranging from 2 kbp to 16 kbp during FIGE and contoured homogeneous electric fields (CHEF). Both supercoiled and linear molecules display a minimum mobility as a function of pulse time in a CHEF apparatus. Linear and cccDNAs of the same size are differently affected by pulse time. Pulse-time dependence was observed for cccDNAs in both systems. Pulse-time dependence in FIGE is very small at a 1.0% agarose concentration, but is pronounced in 0.8% or 1.2% gels.

Full text

PDF
7359

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beverley S. M. Characterization of the 'unusual' mobility of large circular DNAs in pulsed field-gradient electrophoresis. Nucleic Acids Res. 1988 Feb 11;16(3):925–939. doi: 10.1093/nar/16.3.925. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. 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]
  3. Carle G. F., Olson M. V. Separation of chromosomal DNA molecules from yeast by orthogonal-field-alternation gel electrophoresis. Nucleic Acids Res. 1984 Jul 25;12(14):5647–5664. doi: 10.1093/nar/12.14.5647. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chu G., Vollrath D., Davis R. W. Separation of large DNA molecules by contour-clamped homogeneous electric fields. Science. 1986 Dec 19;234(4783):1582–1585. doi: 10.1126/science.3538420. [DOI] [PubMed] [Google Scholar]
  5. Dingman C. W., Fisher M. P., Kakefuda T. Role of molecular conformation in determining the electrophoretic properties of polynucleotides in agarose-acrylamide gels. II. Biochemistry. 1972 Mar 28;11(7):1242–1250. doi: 10.1021/bi00757a020. [DOI] [PubMed] [Google Scholar]
  6. Duggleby R. G., Kinns H., Rood J. I. A computer program for determining the size of DNA restriction fragments. Anal Biochem. 1981 Jan 1;110(1):49–55. doi: 10.1016/0003-2697(81)90110-x. [DOI] [PubMed] [Google Scholar]
  7. FERGUSON K. A. STARCH-GEL ELECTROPHORESIS--APPLICATION TO THE CLASSIFICATION OF PITUITARY PROTEINS AND POLYPEPTIDES. Metabolism. 1964 Oct;13:SUPPL–SUPPL1002. doi: 10.1016/s0026-0495(64)80018-4. [DOI] [PubMed] [Google Scholar]
  8. Hervet H., Bean C. P. Electrophoretic mobility of lambda phage HIND III and HAE III DNA fragments in agarose gels: a detailed study. Biopolymers. 1987 May;26(5):727–742. doi: 10.1002/bip.360260512. [DOI] [PubMed] [Google Scholar]
  9. Hightower R. C., Metge D. W., Santi D. V. Plasmid migration using orthogonal-field-alternation gel electrophoresis. Nucleic Acids Res. 1987 Oct 26;15(20):8387–8398. doi: 10.1093/nar/15.20.8387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Johnson P. H., Grossman L. I. Electrophoresis of DNA in agarose gels. Optimizing separations of conformational isomers of double- and single-stranded DNAs. Biochemistry. 1977 Sep 20;16(19):4217–4225. doi: 10.1021/bi00638a014. [DOI] [PubMed] [Google Scholar]
  11. Johnson P. H., Miller M. J., Grossman L. I. Electrophoresis of DNA in agarose gels. II. Effects of loading mass and electroendosmosis on electrophoretic mobilities. Anal Biochem. 1980 Feb;102(1):159–162. doi: 10.1016/0003-2697(80)90332-2. [DOI] [PubMed] [Google Scholar]
  12. Levene S. D., Zimm B. H. Separations of open-circular DNA using pulsed-field electrophoresis. Proc Natl Acad Sci U S A. 1987 Jun;84(12):4054–4057. doi: 10.1073/pnas.84.12.4054. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lumpkin O. J., Déjardin P., Zimm B. H. Theory of gel electrophoresis of DNA. Biopolymers. 1985 Aug;24(8):1573–1593. doi: 10.1002/bip.360240812. [DOI] [PubMed] [Google Scholar]
  14. Mathew M. K., Hui C. F., Smith C. L., Cantor C. R. High-resolution separation and accurate size determination in pulsed-field gel electrophoresis of DNA. 4. Influence of DNA topology. Biochemistry. 1988 Dec 27;27(26):9222–9226. doi: 10.1021/bi00426a022. [DOI] [PubMed] [Google Scholar]
  15. Mathew M. K., Smith C. L., Cantor C. R. High-resolution separation and accurate size determination in pulsed-field gel electrophoresis of DNA. 1. DNA size standards and the effect of agarose and temperature. Biochemistry. 1988 Dec 27;27(26):9204–9210. doi: 10.1021/bi00426a019. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Mickel S., Arena V., Jr, Bauer W. Physical properties and gel electrophoresis behavior of R12-derived plasmid DNAs. Nucleic Acids Res. 1977;4(5):1465–1482. doi: 10.1093/nar/4.5.1465. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Roode D., Liebschutz R., Maulik S., Friedemann T., Benton D., Kristofferson D. New developments at BIONET. Nucleic Acids Res. 1988 Mar 11;16(5):1857–1859. doi: 10.1093/nar/16.5.1857. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. 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]
  20. Serwer P., Allen J. L. Conformation of double-stranded DNA during agarose gel electrophoresis: fractionation of linear and circular molecules with molecular weights between 3 X 10(6) and 26 X 10(6). Biochemistry. 1984 Feb 28;23(5):922–927. doi: 10.1021/bi00300a020. [DOI] [PubMed] [Google Scholar]
  21. Serwer P. Electrophoresis of duplex deoxyribonucleic acid in multiple-concentration agarose gels: fractionation of molecules with molecular weights between 2 X 10(6) and 110 X 10(6). Biochemistry. 1980 Jun 24;19(13):3001–3004. doi: 10.1021/bi00554a026. [DOI] [PubMed] [Google Scholar]
  22. Serwer P. The mechanism of DNA's fractionation during pulsed-field agarose gel electrophoresis: a hypothesis. Appl Theor Electrophor. 1988;1(1):19–22. [PubMed] [Google Scholar]
  23. Southern E. M. Measurement of DNA length by gel electrophoresis. Anal Biochem. 1979 Dec;100(2):319–323. doi: 10.1016/0003-2697(79)90235-5. [DOI] [PubMed] [Google Scholar]
  24. Stellwagen N. C. Effect of the electric field on the apparent mobility of large DNA fragments in agarose gels. Biopolymers. 1985 Dec;24(12):2243–2255. doi: 10.1002/bip.360241207. [DOI] [PubMed] [Google Scholar]
  25. West R. The electrophoretic mobility of DNA in agarose gel as a function of temperature. Biopolymers. 1987 May;26(5):607–608. doi: 10.1002/bip.360260502. [DOI] [PubMed] [Google Scholar]
  26. Willis C. E., Willis D. G., Holmquist G. P. An equation for DNA electrophoretic mobility in agarose gels. Appl Theor Electrophor. 1988;1(1):11–18. [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES