The effects of different donor cells and passages on development of reconstructed embryos.
In order to study the effects of different donor cells and passages on development of nuclear transfer embryos, we constructed embryos by electrofusing several kinds of donor cells into enucleated M II oocytes from Kun Ming (KM) mouse. These cells include 2-cell embryonic blastomeres, KMW embryonic stem (ES) cells, fetal fibroblast, ear fibroblast, tail tip fibroblast, sertoil cells and spermatogonia. Meanwhile, we compared the effects of passage numbers of fetal fibroblast cells on developmental competency after nuclear transfer. We found that 7.4% of reconstructed embryos from 2-cell embryonic blastomeres and 0.7% from ES cell could develop to blastocyst in vitro; embryos from fetal fibroblast could only develop to morula stage with the rate of 0.2%; embryos from spermatogonia could only develop to 8-cell stage and the rate was 0.3%; embryos respectively from ear fibroblast, sertoli cell and tail tip fibroblast could only develop to 4-cell stage. Although 2-cell development rate of embryos reconstructed from fetal fibroblast in first passage was significantly lower than those from the 2nd, the 3rd and the 4th passage, embryos from different passages could develop to 8-cell stage except the 3rd passage. The result indicated that it is more difficult for terminally differentiated cell nuclei to be reprogrammed in enucleated M II oocytes than for low differentiated cell nuclei. The reason of low development rate from ES cells maybe that most of ES cells was at S stage of the cell cycle, which out of coordination with M II oocytes. We could conclude that culture and passage of donor cells might be benefit to nucleus reprogramming.
Lei
Lei
L
State Key Laboratory of Reproductive Biology, Institute of Zoology, the Chinese Academy of Sciences, Beijing 100080, China.
Liu
Zhong-Hua
ZH
Wang
Hong
H
Kou
Zhao-Hui
ZH
Wu
Yu-Qi
YQ
Xu
Ying
Y
Cheng
Yong
Y
Zhu
Zi-Yu
ZY
Xia
Guo-Liang
GL
Chen
Da-Yuan
DY
D000818:Animals; D002453:Cell Cycle; D002455:Cell Division; D002478:Cells, Cultured; D019976:Cloning, Organism / Q000379:methods*; D004622:Embryo, Mammalian / Q000166:cytology*; D005314:Embryonic and Fetal Development; D005260:Female; D005347:Fibroblasts / Q000166:cytology; D005434:Flow Cytometry; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000166:cytology; D013234:Stem Cells / Q000166:cytology