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KMID : 0359319930330010151
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Korean Journal of Veterinary Research
1993 Volume.33 No. 1 p.151 ~ p.169
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Studies on production of nuclear transplanted mouse embryos
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Abstract
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The present study was carried out to investigate the best condition for nuclear-cytoplasm fusion and in vitro culture of nuclear transplanted embryos and to investigate the production of nuclear transplanted offsprings.
The nuclei from 2-, 4- and 8-cell mouse embryos were transferred into enucleated 2-cell embryos, and the reconstituted embryos were submitted to direct current(DC) pulses at output voltage of 1.0, 1.5 and 2.0㎸/㎝ for 100, 150 and 200μsec to induce cell fusion.
l. The culture of intact or zona cut 2-cell embryos in the medium supplemented with cytochalasin B(5㎍/㎖) and colcemide(0.1㎍/㎖)for 30 and 60 minutes did not affect the development to later stage.
2. The in vitro developmental rates of group A (a nucleus from one of the blastomeres was removed) and B(electrofusion of group A) were significantly lower than that of control group(p<0.01).
3. When nuclear transplanted embryos were submitted to electrofusion, the significantly higher fusion rates of 2-cell donor nuclei were achieved at the electric field strength of DC 1.5㎸/㎝ for 100 and 150μsec, DC 2.0㎸/㎝ for 100∼200μsec than DC 1.0㎸/㎝ for 100 and 150μsec(p<0.01). The significantly higher fusion rates of 4-cell donor nuclei were achieved at DC 2.0㎸/㎝ for 100 and 150μsec than DC 1.0㎸/㎝ for 100∼200μsec(p<0.01). These fusion rates in.8-cell donor nuclei were 88.7∼99.3% .
4. The developmental potency to blastocyst in 2- and 4-cell donor nuclei was significantly higher in DC 1.0 and 2.0㎸/㎝ for 100∼200μsec treated group and DC 2.0㎸/㎝ for 150 and 200μsec treated group (p<0.01) . The developmental potency to blastocyst in 8-cell donor nuclei was significantly higher in DC 2.0㎸/㎝ for 100μsec treated group than in DC 1.0㎸/㎝ for 100μsec treated group and DC 2.0㎸/㎝ for 150 and 200μsec treated group(p<001).
5. The developmental potency to blastocyst after nuclear transplantation was significantly higher in 2-cell donor nuclei than in 8-cell donor nuclei(p<0.01).
6. The success rate of nuclear injection into enucleated 2-cell embryos was significantly higher in 2-cell donor nuclei than in 4- or 8-cell donor nuclei(p<0.01).
7. The culture time taken for the nuclear transplanted 2-cell embryos to blastocyst stage was significantly longer in 2-cell donor nuclei than in 8-cell donor nuclei(p<0.01).
8. There was no significant difference in the developmental potency of nuclear transplanted embryos with in the concentration of EGF at 0 to 15ng per ㎖ of BMOC-3 solution.
9. The production rates of offspring after transfer of nuclear transplanted embryos to recipient mouse were significantly higher in 2-cell donor nuclei than in 8-cell donor nuclei(p<0.01).
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KEYWORD
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