International Mammalian Genome Society

The 16th International Mouse Genome Conference (2002)


POSTER 172 - INFLUENCE OF GENETIC BACKGROUND ON THE CELLULAR CONTRIBUTION TO, AND SUCCESS OF, AGGREGATION CHIMAERAS

M Legge
University of Otago

1) Fox J, 2) Eccles MR
1) Department of Biochemistry 2) Department of Pathology, University of Otago

Aggregation chimaera mice may be used to study the role of mutant genes where the mutation results in tissue agenesis. However, the genetic background of the embryos (mutant and wildtype) may influence the successful generation of aggregation chimaeras. We used mouse embryo aggregation chimaeras to investigate kidney organogenesis, and observe genotype dependent delays in early embryo development. We report here an in-vitro technique we developed to monitor embryo contribution to the aggregation chimaeras. Eight-cell embryos were obtained from C3H/HeJ and C57BL/6J mice. Zona pellucida free embryos from the two inbred strains of mice were aggregated and aggregation confirmed by microscopic analysis.Although both inbred strains achieved the eight-cell stage, the aggregation rate was slow resulting in irregular shaped chimaeras. Embryo culture identified that C3H/HeJ embryos were approximately six-hours behind in development compared to C57BL/6J embryos. Ultimately all irregular shaped aggregations failed to develop. To establish the relative contributions of both genotypes to the aggregation chimaeras we developed the use of PKH67(green) and PKH26(red) cell membrane markers to facilitate the tracking of each embryo during aggregation. These markers demonstrated that the C3H/HeJ contribution to the aggregation was minimal.We conclude that the genetic background of the mice is critical for success, and that the developmental programme of the embryos is an important consideration for aggregation chimaeras. The technique we describe permits a direct, non-invasive, method for the assessment of chimaera formation and a measure of success prior to embryo transfer.


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