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Electrophoretic DNA analysis for the detection of apoptosis

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Abstract

There are many techniques available for the detection of apoptotic cells; some are based on morphological changes, others on biochemical events. However, electrophoretic detection of the systematic cleavage of DNA into oligonucleosomal multimers of 180–200 bp remains the “hallmark” of apoptosis. Conventional constant field agarose gel electrophoresis of DNA from apoptotic cells can be used to resolve the multimers into the characteristic DNA ladders indicative of apoptotic cell death. More recently, it has become clear that the generation of the lower molecular weight oligonucleosomal DNA is preceded by the generation of higher molecular weight fragments. In some cell types, DNA cleavage proceeds no further than the formation of 300 and/or 50 kbp cleavage products. DNA fragmentation of this size can only be resolved using a form of pulsed-field gel electrophoresis. Basic “starter” protocols for conventional and pulsed field electrophoresis for the detection of apoptotic cell DNA are described in this article.

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References

  1. Martin, S. J., Lennoru, S. V., Bonham, A. M., and Cotter, T. G. (1990) Induction of apoptosis (programmed cell death) in human leukaemic HL-60 cells by inhibition of RNA or protein synthesis.J Immunol. 145, 1859–1867.

    CAS  Google Scholar 

  2. Schwartz, D.C. and Cantor, C. R. (1984) Separation of yeast chromosome-sized DNAs by pulsed field gradient gel electrophoresis.Cell 37, 67–75.

    Article  CAS  Google Scholar 

  3. Oberhammer, F., Wilson, J. W., Dive, C., Morris, I. D., Hickman, J. A., Wakeling, A. E., Walker, P. R.. and Sikorska, M. (1993) Apoptotic death in epithelial cells: cleavage of DN A to 300 and/or 50kb fragments prior to or in the absence of internucleosomal fragmentation.EMBO J. 12, 3679–3684.

    CAS  Google Scholar 

  4. Filipski, J., Leblanc, J., Youdale, A., Sikorska, M., and Walker, P. R. (1990) Periodicity of DNA folding in higher order chromatin structures.EMBO J. 9, 1319–1327.

    CAS  Google Scholar 

  5. Brown, D. G., Sun, X.-M., and Cohen, G. M. (1993) Dexamethasone-induced apoptosis involves cleavage of DNA to large fragments prior to internucleosomal fragmentation.J. Biol. Chem. 268, 3037–3039.

    CAS  Google Scholar 

  6. Wood, A. C., Waters, C. M., Garner, A., and Hickman, J. A. (1994) Changes in c-myc expresssion and the kinetics of dexamethasone-induced programmed cell death (apoptosis) in human lymphoid leukaemia cells.Br. J. Cancer 69, 663–669.

    CAS  Google Scholar 

  7. Walker, P. R., Smith, C., Youdale, A., Leblanc, J., Whitfield, J. F., and Sikorska, M. (1991) Topoisomerase II-reactive chemotherapeutic drugs induce apoptosis in thymocytes.Cancer Res. 51, 1078–1085.

    CAS  Google Scholar 

  8. Luo, Y. and Kessel, D. (1996) Detection of early and late stage apoptosis with field inversion gel electrophoresis.Biotechniques 21, 812–816.

    CAS  Google Scholar 

  9. Walker, P. R., Kokileva, L., Leblanc, J., and Sikorska, M. (1993) Detection of the initial stages of DNA fragmentation in apoptosis.BioTechniques 15, 1032–1040.

    CAS  Google Scholar 

Further Reading

  1. Kerr, J. F. R., Winterford, C M., and Harmon, B. V. (1994) Apoptosis: its significance in cancer and cancer therapy.Cancer 73, 2013–2026.

    Article  CAS  Google Scholar 

  2. Levine, A. J. (1997) p53, the cellular gatekeeper for growth and division.Cell 88, 323–331.

    Article  CAS  Google Scholar 

  3. Yang, E. and Korsmeyer, S. J. (1996) Molecular thanotopsis: a discourse on the BCL-2 family and cell death.Blood 88, 386–401.

    CAS  Google Scholar 

  4. Smith, C. L. (ed.) (1993) Paper Symposium: Changing directions in electrophoresis.Electrophoresis 14, 249 – 370.

    Google Scholar 

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Correspondence to Paul D. Allen.

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Allen, P.D., Newland, A.C. Electrophoretic DNA analysis for the detection of apoptosis. Mol Biotechnol 9, 247–251 (1998). https://doi.org/10.1007/BF02915798

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  • DOI: https://doi.org/10.1007/BF02915798

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