TY - JOUR

T1 - Analysis of nucleolar morphology and protein localization as an indicator of nuclear reprogramming

AU - Østrup, Olga

AU - Pedersen, Hanne Skovsgaard

AU - Holm, Hanne M.

AU - Hyttel, Poul

PY - 2015

Y1 - 2015

N2 - 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.

AB - 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.

U2 - 10.1007/978-1-4939-1594-1_12

DO - 10.1007/978-1-4939-1594-1_12

M3 - Journal article

C2 - 25287345

VL - 1222

SP - 161

EP - 174

JO - Methods in Molecular Biology

JF - Methods in Molecular Biology

SN - 1064-3745

ER -