A simple method for producing tetraploid porcine parthenogenetic embryos

The objective was to produce porcine tetraploid parthenogenetic embryos using cytochalasin B, which inhibits polar body extrusion. Porcine cumulus-enclosed oocytes aspirated from antral follicles were cultured for 51 h, and treated with cytochalasin B from 35 h to 42 h after the start of culture. After maturation culture, 74.7% (2074/2775) of oocytes treated with cytochalasin B did not extrude a polar body (OPB oocytes). In contrast, 80.4% (1931/2403) of control oocytes extruded a polar body (1PB oocytes). The OPB oocytes were electrically stimulated, treated with cytochalasin B again for 3 h, and then cultured without cytochalasin B. Six days after electrical stimulation, 49.8% (321/644) reached the blastocyst stage. The number of cells in these blastocysts derived from OPB oocytes was significantly lower than that from 1PB oocytes (OPB: 24.9 +/- 10.6; 1PB: 43.0 +/- 17.1; mean +/- SD). A porcine chromosome 1-specific sequence was detected in parthenogenetic OPB embryos by fluorescence in situ hybridization (FISH) analysis. Typical pronucleus-stage samples derived from OPB embryos had two pronuclei, each with two signals. In two-cell and blastocyst-stage embryos, four signals were detected in each nucleus derived from OPB embryos. We inferred that OPB oocytes, which had a tetraploid number of chromosomes, started to develop as tetraploid parthenotes after electrical stimulation, and that tetraploid status was stably maintained during early embryonic development, at least until the blastocyst stage. (C) 2011 Elsevier Inc. All rights reserved.