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Pulsed field gel electrophoresis: Theory, instruments and application

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Abstract

Pulsed field gel electrophoresis (PFGE) is a technique for the fractionation of high-molecular-weight DNA ranging from 10 kb to 10 Mb by electrophoresis in agarose gel with an electric field that alternates (pulsates) in two directions. This technology plays a key role in modern genomics, as it allows manipulations with DNA of whole chromosomes or their large fragments. In this review, we discuss (1) the theory behind PFGE; (2) different instruments based on the principle of pulsed field, as well as their advantages and limitations; (3) factors affecting the DNA mobility in PFGE gel; and (4) practical applications of the technique.

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Abbreviations

CHEF:

contour-clamped homogeneous electric field

FIGE:

field inversion gel electrophoresis

OFAGE:

orthogonal field-alternating gel electrophoresis

PFGE:

pulsed field gel electrophoresis or pulsed field gradient electrophoresis (as previously used)

RGE:

rotating gel electrophoresis

TAFE:

transverse alternating field electrophoresis

References

  • Anand, R., Pulsed Field Gel Electrophoresis: A Technique for Fractionating Large DNA Molecules, Trends Genet., 1986, vol. 2, pp. 278–283.

  • Bautsch, W., Bacterial Genome Mapping by Two-Dimensional Pulsed-Field Gel Electrophoresis (2D-PFGE), in Methods in Molecular Biology, Totowa: Humana Press, 1992, vol. 12, pp. 185–201.

    Google Scholar 

  • Birren, B.W., Lai, E., Clark, S.M., Hood, L., and Simon, M.I., Optimized Conditions for Pulsed Field Gel Electrophoretic Separations of DNA, Nucleic Acids Res., 1988, vol. 16, pp. 7563–7582.

    Article  PubMed  CAS  Google Scholar 

  • Britto, C., Ravel, C., Bastien, P., Blaineau, C., Pages, M., Dedet, J.P., and Wincker, P., Conserved Linkage Groups Associated with Large-Scale Chromosomal Rearrangements between Old World and New World Leishmania Genomes, Gene, 1998, vol. 222, pp. 107–117.

    Article  PubMed  CAS  Google Scholar 

  • Burmeister, M. and Lehrach, H., Long-Range Restriction Map around the Duchenne Muscular Dystrophy Gene, Nature, 1986, vol. 324, pp. 582–585.

    Article  PubMed  CAS  Google Scholar 

  • Bustamante, C., Gurrieri, S., and Smith, S.B., Observation of Single DNA Molecules during Pulsed-Field Gel Electrophoresis by Fluorescence Microscopy, in Methods: A Companion to Methods in Enzymology, San Diego: Academic, 1990, vol. 1, pp. 151–159.

    Google Scholar 

  • Cantor, C.R., Gaal, A., and Smith, C.J., High-Resolution Separation and Accurate Size Determination in Pulsed-Field Gel Electrophoresis of DNA. 3. Effect of Electrical Field Shape, Biochemistry, 1988, vol. 27, pp. 9216–9221.

    Article  PubMed  CAS  Google Scholar 

  • Carle, G.F. and Olson, M.V., An Electrophoretic Karyotype for Yeast, Proc. Natl. Acad. Sci. USA, 1985, vol. 82, pp. 3756–3760.

    Article  PubMed  CAS  Google Scholar 

  • Carle, G.F. and Olson, M.V., Separation of Chromosomal DNA Molecules from Yeast by Orthogonal Field Alternation Gel Electrophoresis, Nucleic Acids Res., 1984, vol. 12, pp. 5647–5664.

    Article  PubMed  CAS  Google Scholar 

  • Carle, G.F., Frank, M., and Olson, M.V., Electrophoretic Separation of Large DNA Molecules by Periodic Inversion of the Electric Field, Science, 1986 vol. 232, pp. 65–68.

    Article  PubMed  CAS  Google Scholar 

  • Chu, G. and Gunderson, K., Separation of Large DNA by a Variable-Angle Contour-Clamped Homogeneous Electric Field Apparatus, Anal. Biochem., 1991, vol. 194, pp. 439–446.

    Article  PubMed  CAS  Google Scholar 

  • Chu, G., Pulsed-Field Gel Electrophoresis: Theory and Practice, in Methods: A Companion to Methods in Enzymology, San Diego: Academic Press, 1990, vol. 1, pp. 129–142.

    Google Scholar 

  • Chu, G., Vollrath, D., and Davis, R.W., Separation of Large DNA Molecules by Contour-Clamped Homogeneous Electric Fields, Science, 1986, vol. 234, pp. 1582–1585.

    Article  PubMed  CAS  Google Scholar 

  • Clark, S.M., Lai, E., Birren, B.W., and Hood, L., A Novel Instrument for Separating Large DNA Molecules with Pulsed Homogeneous Electric Fields, Science, 1988, vol. 241, pp. 1203–1205.

    Article  PubMed  CAS  Google Scholar 

  • Cooney, C.A., Separation and Size Determination of DNA over a 10–200 kbp Range, in Methods in Molecular Biology, Totowa: Humana Press, 1992, vol. 12, pp. 31–38.

    Google Scholar 

  • Correia, A., Martin, J.F., and Castro, J.M., Pulsed-Field Gel Electrophoresis Analysis of the Genome of Amino Acid Producing Corynebacteria: Chromosome Sizes and Diversity of Restriction Patterns, Microbiology, 1994, vol. 140, pp. 2841–2847.

    Article  PubMed  CAS  Google Scholar 

  • Dingwall, A. and Shapiro, L., Rate, Origin, and Bidirectionality of Caulobacter Chromosome Replication as Determined by Pulsed-Field Gel Electrophoresis, Proc. Natl. Acad. Sci. USA, 1989, vol. 86, pp. 119–123.

    Article  PubMed  CAS  Google Scholar 

  • Dingwall, A., Shapiro, L., and Ely, B., Analysis of Bacterial Genome Organization and Replication Using Pulsed-Field Gel Electrophoresis, in Methods: A Companion to Methods of Enzymology, San Diego: Academic Press, 1990, vol. 1, pp. 160–168.

    Google Scholar 

  • Dujon, B., Yeasts Illustrate the Molecular Mechanisms of Eukaryotic Genome Evolution, Trends Genet., 2006, vol. 22, pp. 375–387.

    Article  PubMed  CAS  Google Scholar 

  • Ecker, J., PFGE and YAC Analysis of the Arabidopsis Genome, in Methods: A Companion to Methods of Enzymology, San Diego: Academic Press, 1990, vol. 1, pp. 186–194.

    Google Scholar 

  • Eichinger, L., Pachebat, J.A., Glöckner, G., et al, The genome of the Social amoeba Dictyostelium discoideum, Nature, 2005, vol. 435, pp. 43–57.

    Article  PubMed  CAS  Google Scholar 

  • Gardner, M.J., Hall, N., Fung, E., et al, Genome Sequence of the Human Malaria Parasite Plasmodium Falciparum, Nature, 2002, vol. 419, pp. 498–511.

    Article  PubMed  CAS  Google Scholar 

  • Fan, J.B., Chikashige, Y., Smith, C.L., Niwa, O., Yanagida, M., and Cantor, C.R., Construction of a Not I Restriction Map of the Fission Yeast Schizosaccharomyces pombe Genome, Nucleic Acids Res., 1989, vol. 17, pp. 2801–2818.

    Article  PubMed  CAS  Google Scholar 

  • Fischer, G., James, S.A., Roberts, I., Oliver, S., and Louis, E., Chromosomal Evolution in Saccharomyces, Nature, 2000, vol. 405, pp. 451–454.

    Article  PubMed  CAS  Google Scholar 

  • Fischer, G., Rocha, E.P., Brunet, F., Vergassola, M., and Dujon, B., Highly Variable Rates of Genome Rearrangements between Hemiascomycetous Yeast Lineages, PLoS Genet., 2006, vol. 2, pp. 0253–0261.

    Article  CAS  Google Scholar 

  • Fraissinet-Tachet, L., Reymond-Cotton, P., and Fevre, M., Molecular Karyotype of the Phytopathogenic Fungus Sclerotinia sclerotiorum, Curr. Genet., 1996, vol. 29, pp. 496–501.

    Article  PubMed  CAS  Google Scholar 

  • Fujiwara, S., Okuyama, S., and Imanaka, T., The World of Archaea: Genome Analysis, Evolution and Thermostable Enzymes, Gene, 1996, vol. 179, pp. 165–170.

    Article  PubMed  CAS  Google Scholar 

  • Gardiner, K., Laas, W., and Patterson, D., Fractionation of Large Mammalian DNA Restriction Fragments using Vertical Pulsed-Field Gradient Gel Electrophoresis, Somatic Cell Molec. Genet., 1986, vol. 12, pp. 185–195.

    Article  CAS  Google Scholar 

  • Gardiner, K., Transverse Alternating-Field Electrophoresis, in Methods in Molecular Biology, Totowa: Humana Press, 1992, vol. 12, pp. 51–61.

    Google Scholar 

  • Giannini, S.H., Schittini, M., Keithly, J.S., Warburton, P., Cantor, C.R., and Van der Ploeg, L.H.T., Karyotype Analysis of Leishmania Species and Its Use in Classification and Clinical Diagnosis, Science, 1986, vol. 232, pp. 761–765.

    Article  Google Scholar 

  • Gunderson, K. and Chu, G., Pulsed-Field Electrophoresis of Multimegabase-Sized DNA, Mol. Cell. Biol., 1991, vol. 11, pp. 3348–3354.

    PubMed  CAS  Google Scholar 

  • Gurrieri, S., Smith, S., and Bustamante, C., Trapping of Megabase-Sized DNA Molecules during Agarose Gel Electrophoresis, Proc. Natl. Acad. Sci. USA, 1999, vol. 96, pp. 453–458.

    Article  PubMed  CAS  Google Scholar 

  • Gurrieri, S., Smith, S.B., Wells, K.S., Johnson, I.D., and Bustamante, C., Real-Time Imaging of the Reorientation Mechanisms of YOYO-Labelled DNA Molecules during 90° and 120° Pulsed Field Gel Electrophoresis, Nucleic Acids Res., 1996, vol. 24, pp. 4759–4767.

    Article  PubMed  CAS  Google Scholar 

  • Henriksson, J., Dujardin, J.C., Barnabe, C., Brisse, S., Timperman, G., Venegas, J., Pettersson, U., Tibayrenc, M., and Solari, A., Chromosomal Size Variation in Trypanosoma cruzi Is Mainly Progressive and Is Evolutionarily Informative, Parasitology, 2002, vol. 124, pp. 277–286.

    Article  PubMed  CAS  Google Scholar 

  • Henriksson, J., Porcel, B., Rydaker, M., Ruiz, A., Sabaj, V., Galanti, N., Cazzulo, J.J., Frasch, A.C., and Pettersson, U., Chromosome Specific Markers Reveal Conserved Linkage Groups in Spite of Extensive Chromosomal Size Variation in Trypanosoma cruzi, Mol. Biochem. Parasitol., 1995, vol. 73, pp. 63–74.

    Article  PubMed  CAS  Google Scholar 

  • Kuspa, A., Vollrath, D., Cheng, Y., and Kaiser, D., Physical Mapping of the Myxococcus xanthus Genome by Random Cloning in Yeast Artificial Chromosomes, Proc. Natl. Acad. Sci. USA, 1989, vol. 86, pp. 8917–8921.

    Article  PubMed  CAS  Google Scholar 

  • Lai, E., Birren, B.W., Clark, S.M., Simon, M.I., and Hood, L., Pulsed Field Gel Electrophoresis, Biotechniques, 1989, vol. 7, pp. 34–42.

    PubMed  CAS  Google Scholar 

  • Larin, Z., Functional Analysis of Mammalian Genomes using Yeast Artificial Chromosomes, in Pulsed Field Gel Electrophoresis: A Practical Approach, New York: IRL Press at Oxford University Press Inc., 1995, 139–157.

    Google Scholar 

  • Mathew, M.K., Smith, C.J., and Cantor, C.R., 1988b. High-Resolution Separation and Accurate Size Determination in Pulsed-Field Gel Electrophoresis of DNA. 2. Effect of Pulse Time and Electric Field Strength and Implications for Models of the Separation Process, Biochemistry, 1988b, vol. 27, pp. 9210–9216.

    Article  PubMed  CAS  Google Scholar 

  • Mathew, M.K., Smith, C.J., and 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, 1988a, vol. 27, pp. 9204–9210.

    Article  PubMed  CAS  Google Scholar 

  • Mezhevaya, E.V., Stepanova, V.P., and Yarovoi, B.F., Pulsed-Field Gel Electrophoresis as a New Tool in Genetics, in Uspekhi sovremennoi genetiki (Advances in Modern Genetics), Moscow: Nauka, 1993, vol. 18, pp. 136–162.

    Google Scholar 

  • Ohki, M. and Smith, C.L., Tracking Bacterial DNA Replication Forks in Vivo by Pulsed Field Gel Electrophoresis, Nucleic Acids Res., 1989, vol. 17, pp. 3479–3490.

    Article  PubMed  CAS  Google Scholar 

  • Orbach, M.J., Vollrath, D., Davis, R.W., and Janofsky, C., An Electrophoretic Karyotype of Neurospora crassa, Mol. Cell. Biol., 1988, vol. 8, pp. 1469–1473.

    PubMed  CAS  Google Scholar 

  • Ougen, P. and Cohen, D., Yeast Artificial Chromosome Cloning using PFGE, in Pulsed Field Gel Electrophoresis: A Practical Approach, New York: IRL Press at Oxford University Press Inc., 1995, pp. 95–118.

    Google Scholar 

  • Pyle, L.E. and Finch, L.R., A Physical Map of the Genome of Mycoplasma mycoides Subspecies Mycoides Y with Some Functional Loci, Nucleic Acids Res., 1988, vol. 16, pp. 6027–6039.

    Article  PubMed  CAS  Google Scholar 

  • Ragoussis, J., PFGE Analysis of Yeast Artificial Chromosomes, in Pulsed Field Gel Electrophoresis: A Practical Approach, New York: IRL Press at Oxford University Press Inc., 1995, pp. 119–137.

    Google Scholar 

  • Schwartz, D.C. and Cantor, C.R., Separation of Yeast Chromosome-Sized DNAs by Pulsed Field Gradient Gel Electrophoresis, Cell, 1984, vol. 37, pp. 67–75.

    Article  PubMed  CAS  Google Scholar 

  • Schwartz, D.C., Saffran, W., Welsh, J., Haas, R., Goldenberg, M., and Cantor, C.R., New Techniques for Purifying Large DNAs and Studying Their Properties and Packaging, Cold Spring Harbor Symp. Quant. Biol., 1982, vol. 47, pp. 189–195.

    CAS  Google Scholar 

  • Smith, C.L. and Condemine, G., New Approaches for Physical Mapping of Small Genomes, J. Bacteriol., 1990, vol. 172, pp. 1167–1172.

    PubMed  CAS  Google Scholar 

  • Smith, C.L., Econome, J.G., Schutt, A., Klco, S., and Cantor, C.R., A Physical Map of the Escherichia coli K12 Genome, Science, 1987, vol. 236, pp. 1448–1453.

    Article  PubMed  CAS  Google Scholar 

  • Smith, N., Klco, S.R., and Cantor, Ch.R., Pulsed-Field Gel Electrophoresis and Methods of Work with Large DNA Molecules, in Genome Analysis: A Practical Approach, Davies, K.E., Ed., Oxford: IRL Press, 1988. Translated under the title Analiz genoma. Metody, Moscow: Mir, 1990, pp. 58–94.

    Google Scholar 

  • Southern, E.M. and Elder, J.K., Theories of Gel Electrophoresis of High Molecular Weight DNA, in Pulsed Field Gel Electrophoresis: A Practical Approach, New York: IRL Press at Oxford University Press Inc., 1995, pp. 1–19.

    Google Scholar 

  • Southern, E.M., Anand, R., Brown, W.R.A., and Fletcher, D.S., A Model for the Separation of Large DNA Molecules by Crossed Field Gel Electrophoresis, Nucleic Acids Res., 1987, vol. 15, pp. 5925–5943.

    Article  PubMed  CAS  Google Scholar 

  • Steward, G.F., Fingerprinting Viral Assemblages by Pulsed Field Gel Electrophoresis (PFGE), in Marine Microbiology. Methods in Microbiology Series, London: Academic Press, 2001, vol. 30, pp. 85–103.

    Google Scholar 

  • Taylor, J.L., Borgmann, I., and Seguin-Swartz, G., Electrophoretic Karyotyping of Leptosphaeria maculans Differentiates Highly Virulent from Weakly Virulent Isolates, Curr. Genet., 1991, vol. 19, pp. 273–277.

    Article  CAS  Google Scholar 

  • Taylor, J.W., Geiser, D.M., Burt, A., and Koufopanou, V., The Evolutionary Biology and Population Genetics Underlying Fungal Strain Typing, Clin. Microbiol. Rev., 1999, vol. 12, pp. 126–146.

    PubMed  CAS  Google Scholar 

  • Ziegler, A., Geiger, K.H., Ragoussis, J., and Szalay, G., A New Electrophoresis Apparatus for Separating Very Large DNA Molecules, J. Clin. Chem. Clin. Biochem., 1987, vol. 25, pp. 578–579.

    Google Scholar 

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  1. Institute of Cytology, Russian Academy of Sciences, St. Petersburg, Russia

    E. S. Nassonova

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Original Russian Text © E.S. Nassonova, 2008, published in Tsitologiya, Vol. 50, no. 6, 2008.

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Nassonova, E.S. Pulsed field gel electrophoresis: Theory, instruments and application. Cell Tiss. Biol. 2, 557–565 (2008). https://doi.org/10.1134/S1990519X08060011

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