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Analysis of Nucleolar Morphology and Protein Localization as an Indicator of Nuclear Reprogramming

  • Olga Østrup
  • Hanne S. Pedersen
  • Hanne M. Holm
  • Poul HyttelEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1222)

Abstract

When a cell is reprogrammed to a new phenotype, the nucleolus undergoes more or less dramatic modulations, which can be used as a marker for the occurrence of the reprogramming. This phenomenon is most pronounced when differentiated cells are reprogrammed to totipotency when they are submitted to cloning by somatic cell nuclear transfer. However, when cells are reprogrammed by less fundamental means, as for example treatment by Xenopus extract or expression of pluripotency genes, more subtle nucleolar modulations can also be noted. The monitoring and understanding of the reprogramming-related nucleolar modulations are based upon detailed knowledge about the nucleolar changes that occur during normal development from the developing oocyte over oocyte maturation and fertilization to the activation of the embryonic genome in the early embryo. Below, the ultrastructural and molecular modulations of the nucleolus are summarized in this developmental context, but also as they occur in assisted reproductive technologies such as in vitro fertilization and somatic cell nuclear transfer. Moreover, detailed protocols for monitoring the nucleolar changes by transmission electron microscopy and immunocytochemistry are presented.

Key words

Nucleolus Nucleus Transmission electron microscopy Immunocytochemistry Assisted reproductive technology 

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Olga Østrup
    • 1
  • Hanne S. Pedersen
    • 2
  • Hanne M. Holm
    • 3
  • Poul Hyttel
    • 3
    Email author
  1. 1.Department of Tumor BiologyOslo University Hospital RHOsloNorway
  2. 2.Department of Animal ScienceAarhus UniversityAarhusDenmark
  3. 3.Department of Clinical Veterinary and Animal SciencesUniversity of CopenhagenFrederiksbergDenmark

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