Elsevier

Methods in Enzymology

Volume 96, 1983, Pages 597-608
Methods in Enzymology

[48] Proteins of pore complex-lamina structures from nuclei and nuclear membranes

https://doi.org/10.1016/S0076-6879(83)96052-4Get rights and content

Publisher Summary

The nuclear envelope (NE) is composed of three prominent architectural components: the inner and outer nuclear membrane and the pore walls, the nonmembranous pore complex material, and the nuclear lamina. This chapter describes some simple procedures for the isolation of pore complex-lamina structures, such as procedure for isolation of nuclear envelopes from Xenopus laevis oocytes that involves preparation of nuclear envelopes from manually isolated oocyte nuclei, and large-scale isolation of nuclear envelopes from mass-isolated oocyte nuclei. The architectural components of the nuclear pore complexes and the nuclear lamina that interconnects them are the only NE structures resistant to treatments with nucleases, low- and high-salt buffers, and nonionic detergents. The purest pore complex-lamina fractions are obtained, when nuclear membranes or whole nuclei are extracted, after removal of chromatin, with buffers containing high salt concentrations and nonionic detergents. This nucleocortical skeleton structure cannot be enriched in comparable purity and structural preservation by other approaches—for example, using buffers containing urea, guanidinium hydrochloride, heparin, or sodium deoxycholate for extraction. The other methods presented are for isolation of nuclear pore complex-lamina structures from erythrocytes of Xenopus laevis and of chicken, and nuclear pore complex-lamina structures from nuclear membranes of rat liver.

References (23)

  • S. Ely et al.

    Exp. Cell Res.

    (1978)
  • G. Krohne et al.

    Exp. Cell Res.

    (1978)
  • K.R. Shelton et al.

    J. Biol. Chem.

    (1980)
  • G. Krohne et al.

    J. Mol. Biol.

    (1981)
  • R. Stick et al.

    Exp. Cell Res.

    (1982)
  • G.G. Maul et al.

    Exp. Cell Res.

    (1980)
  • J.G. Gall

    Methods Cell Physiol.

    (1966)
  • S.H. Kaufmann et al.

    J. Biol. Chem.

    (1983)
  • W.W. Franke

    Int. Rev. Cytol., Suppl.

    (1974)
  • G.G. Maul

    Int. Rev. Cytol., Suppl.

    (1977)
  • D.W. Fawcett

    Am. J. Anat.

    (1966)
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      If the preparation is scaled down, volumes of buffers should be reduced proportionally. The preparation of nuclei is a combination of the methods described by Krohne and Franke (1983) and Kaufmann et al. (1983). All preparation should be carried out at 4 °C if not indicated otherwise, and the solutions contain a final concentration of 10 μg trypsin inhibitor⧸ml and 0.2 mM PMSF⧸ml.

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