摘要
The aim of this study was to optimize electrofusion conditions for generating porcine tet- raploid （4n） embryos and produce tetraploid/diploid （4n/2n） chimeric embryos. Different electric field intensities were tested and 2 direct current （DC） pulses of 0.9 kV/em for 30 p.s was selected as the optimum condition for electrofusion of 2-cell embryos to produce 4n embryos. The fusion rate of 2-cell embryos and the development rate to blastocyst of presumably 4n embryos, reached 85.4% and 28.5%, respectively. 68.18% of the fused embryos were found to be 4n as demonstrated by fluorescent in situ hybridization （FISH）. Although the number of blastomeres in 4n blastocysts was significantly lower than in 2n blastocysts （P 〈 0.05）, there was no significant difference in developmental rates of blastocysts between 2n and 4n embryos （P 〉 0.05）, suggesting that the blas- tocyst forming capacity in 4n embryos is similar to those in 2n embryos. Moreover, 4n/2n chimeric embryos were obtained by aggregation of 4n and 2n embryos. We found that the developmental rate and cell number of blastocysts of 4-cell （4n）/4-cell （2n） chimeric embryos were significantly higher than those of 2-cell （4n）/4-cell （2n）, 4-cell （4n）/8-cell （2n）, 4-cell （4n）/2-cell （2n） chimeric embryos （P 〈 0.05）. Consistent with mouse chimeras, the majority of 4n cells contribute to the trophectoderm （TE）, while the 2n cells are mainly present in the inner cell mass （ICM） of porcine 4n/2n chimeric embryos. Our study established a feasible and efficient approach to produce porcine 4n embryos and 4n/2n chimeric embryos.
The aim of this study was to optimize electrofusion conditions for generating porcine tet- raploid （4n） embryos and produce tetraploid/diploid （4n/2n） chimeric embryos. Different electric field intensities were tested and 2 direct current （DC） pulses of 0.9 kV/em for 30 p.s was selected as the optimum condition for electrofusion of 2-cell embryos to produce 4n embryos. The fusion rate of 2-cell embryos and the development rate to blastocyst of presumably 4n embryos, reached 85.4% and 28.5%, respectively. 68.18% of the fused embryos were found to be 4n as demonstrated by fluorescent in situ hybridization （FISH）. Although the number of blastomeres in 4n blastocysts was significantly lower than in 2n blastocysts （P 〈 0.05）, there was no significant difference in developmental rates of blastocysts between 2n and 4n embryos （P 〉 0.05）, suggesting that the blas- tocyst forming capacity in 4n embryos is similar to those in 2n embryos. Moreover, 4n/2n chimeric embryos were obtained by aggregation of 4n and 2n embryos. We found that the developmental rate and cell number of blastocysts of 4-cell （4n）/4-cell （2n） chimeric embryos were significantly higher than those of 2-cell （4n）/4-cell （2n）, 4-cell （4n）/8-cell （2n）, 4-cell （4n）/2-cell （2n） chimeric embryos （P 〈 0.05）. Consistent with mouse chimeras, the majority of 4n cells contribute to the trophectoderm （TE）, while the 2n cells are mainly present in the inner cell mass （ICM） of porcine 4n/2n chimeric embryos. Our study established a feasible and efficient approach to produce porcine 4n embryos and 4n/2n chimeric embryos.
基金
supported by grants from the National Basic Research Program of China(973 Program
Grant No.2009CB941000 and 2011CB944202)
the Fund for Outstanding Young Scholars in Heilongjiang Province(Grant No.JC200905)
