Gene Knockout Protocols

Volume 158 of the series Methods in Molecular Biology pp 135-154

Aggregation Chimeras

Combining ES Cells, Diploid and Tetraploid Embryos
  • Mika TanakaAffiliated withSamuel Lunenfeld Research Institute, Mount Sinai Hospital
  • , Anna-Katerina HadjantonakisAffiliated withSamuel Lunenfeld Research Institute, Mount Sinai Hospital
  • , Andras NagyAffiliated withSamuel Lunenfeld Research Institute, Mount Sinai Hospital

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In recent years, chimeras have been providing a powerful way to study mouse development (1) in combination with invention and improvement of other techniques and materials, including embryonic stem (ES) cells (2) and tetraploid embryos (3,4). ES cells are pluripotent cell lines derived from late blastocyst-stage embryos, which are capable of differentiating into all derivatives of the primitive ectoderm (see Fig. 1) when aggregated with or injected into diploid embryos (5). In contrast, tetraploid embryos, which can be made by electrofusing two cell-stage diploid embryos (3,6,7), have been found to contribute preferentially to most of the extraembryonic cell lineages, i.e., the trophoblast (trophectoderm derivatives) and primitive endoderm derivatives (see Fig. 1) when aggregated with diploid embryos (38). Interestingly, ES cells show a deficiency in extraembryonic lineages, therefore these cells and tetraploid embryo derived cells have a complementary distribution in chimeras made between them. In such chimeras, the embryo proper, the amnion, the yolk sac mesoderm, the allantois and the chorionic mesoderm-derived part of the placenta are completely ES cell-derived, whereas the yolk sac endoderm and the trophoblast cell lineages are tetraploid embryo derived (3,7,9). It is certain that the ES cell⇔tetraploid embryo aggregates have an attractive feature in that they are a reliable and simple way of producing completely ES cell-derived embryos from developmentally competent cell lines (210). This feature is promoting their application in an increasing number of studies.
Fig. 1.

Schematic representation of the various embryo proper and extraembryonic lineages and their relation to each other.