6-DMAP和胚胎干细胞代数对异种重构卵体外发育的影响(简报)

核移植技术已经广泛应用于动物克隆,但是克隆动物的成活率仍然很低。许多克隆胚胎死于妊娠期,少部分能发育到期,正常出生,但多数在出生后由于心肺和消化道的问题,很快就夭折,有些克隆动物有异常表型,如出生时体重和胎盘过大等。研究发现,在同种克隆动物实验中用胚胎干细胞(Embryonic stem cell,ES细胞)作为核供体,发育到期的克隆动物比例明显高于体细胞,并且用杂交一代的小鼠ES细胞为核供体,绝大多数克隆仔

EFFECTS OF 6-DIMETHYLAMINOPURINE AND PASSAGES OF EMBRYONIC STEM CELLS ON THE DEVELOPMENT IN VITRO OF INTERSPECIFIC RECONSTRUCTED OOCYTES

The development of reconstructed cocytes and the survival rate of cloned animal were affected by many factors during nuclear transfer. The genetic constitution and the genetic state of donor nucleus were proposed to be primary factors, which affected the survival rate of cloned animal. In addition, the survival rate of cloned animal might be influenced by nuclear transfer technique itself and passages of donor cells as well as the activation methods of oocytes. We reconstructed oocytes with outbreeding Kunming albino mouse ES cells and enucleated rabbit oocytes, and analyzed the effects of the passages of ES cells and 6-DMAP on the development of interspecific reconstructed cocytes. The interspecific reconstructed ES-ribbit oocytes were activated either by combined two set electric pulses and 6-DMAP or by two set electric pulses alone. The rate of cleavage was significantly higher for the group (86. 2%) treated with 6-DMAP than the group (64.2%,P<0.05) treated with electric pulses only, and the rate of blastocysts was 17.0% and 13.4% respectively, which were not significantly different between two groups. When ES cells that had been passed for 24 and 14 generations were used as donors, the cleavage rates of the reconstructed oocytes were 88. 5% and 82. 1% , respectively (P> 0.05), and the rates of blastolation were 16.7% and 15.4% , respectively (P>0.05). The results show that 6-DMAP increases the cleavage rate of reconstructed oocytes derived from ES cells, and affects slightly the developmental rate of blastocysts. There are no differences when high passage and low passage ES cells are used as nuclear donors.