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Visualization of second polar body chromosomes in fertilized and artificially activated mouse oocytes treated with
A P Dyban1, P De Sutter, D Dozortsev
1Reproductive Genetics Institute, Illinois Masonic Medical Center, Chicago 60657.
Objective:
Our objective was to develop a new reliable method for cytogenetic analysis of the chromosome set in second polar bodies (PBs) from one-cell-stage mouse embryos.
Setting:
The study took place at the Reproductive Biology and Experimental Cytogenetics Laboratories.
Methods:
Oocytes from F1 hybrid and T6/T6 mice were fertilized in vitro and artificially activated with ethanol. Zygotes, parthenogenetic embryos, and isolated second PBs were treated with 10 microM okadaic acid (OA) for 1-2 hr, further cultured in plain medium, and fixed. Chromosomal preparations were made and C-banded, and the number of chromosomes in second PBs and embryos was counted.
Results:
OA-induced nuclear envelope breakdown in pronuclei as well as in second PB nuclei. Countable chromosome plates were obtained in 92-93% of second PBs treated 4-4.5 hr after activation. The T6 marker chromosome could easily be recognized in second PBs from T6/T6 mice. A haploid set of chromosomes was obtained in 18 of 19 isolated second PBs treated with OA 4-5 hr after activation.
Conclusion:
Treatment of second PBs with OA allows visualization of the PB chromosomes. Cytogenetic analysis of the second PB and the corresponding oocyte constitutes a new approach for the study of meiotic nondisjunction in experimental cytogenetics. The chromosomal study of isolated second PBs seems to be promising for clinical preimplantation cytogenetics.