Roscovitine同期化供核细胞对猴—猪异种核移植胚胎发育的影响  

Effects of Roscovitine-treated donor cells on synchronization and development of monkey-porcine inter-species nuclear transplantation embryos

作　　者：许常龙[1] 秦祖兴[1] 梁明明[1] 胡林林[1] 罗茜[1] 崔静辉[1] 罗军[1] 左二伟[1] 许惠艳[1] 卢晟盛[1] 卢克焕[1] 

机构地区：[1]亚热带农业生物资源保护与利用国家重点实验室/广西大学动物繁殖研究所,南宁530005

出　　处：《南方农业学报》2013年第10期1722-1726,共5页Journal of Southern Agriculture

基　　金：国家自然科学基金项目(30860039);广西自然科学基金重点项目(2010GXNSFD013021)

摘　　要：【目的】探讨Roscovitine同期化供核细胞对食蟹猴—猪异种体细胞核移植胚胎体外发育的影响,为提高灵长类的核移植效率奠定基础。【方法】活体采集4岁雄性食蟹猴的耳组织,经组织块培养获得纯化的食蟹猴耳成纤维细胞,细胞经固定、染色后用流式细胞仪分析细胞周期分布。以不同同期化方法处理的食蟹猴耳纤维细胞为供体细胞,以体外成熟、去核的猪卵母细胞为受体细胞,利用电融合法构建食蟹猴—猪异种核移植胚胎,观察异种核移植重构胚胎的体外发育情况。【结果】以15μmol/L Roscovitine处理食蟹猴耳成纤维细胞24、48、72 h获得的G0/G1期细胞比率分别为76.51%、89.69%和90.49%,其中处理48和72 h的G0/G1期细胞比率显著高于对照组(P<0.05)。血清饥饿、接触抑制72 h同期化获得的G0/G1期细胞比率分别为91.12%和90.46%。Roscovitine同期化处理食蟹猴耳成纤维细胞48 h可有效提高食蟹猴—猪异种核移植重构胚胎的囊胚形成率(15.05%),显著高于血清饥饿同期化处理和接触抑制同期化处理的效果(P<0.05)。【结论】Roscovitine可有效同期化食蟹猴耳成纤维细胞在G0/G1期,最终提高食蟹猴—猪异种核移植重构胚胎的囊胚形成率。[Objective]The present study evaluated the effects of Roscovitine on long-tailed macaque ear-derived fi- broblast cell cycle synchronization and embryo development of monkey-porcine interspecies nuclear transfer to provide ref- erences for improving nuclear transfer of primates. [ Method ] Cells were obtained from the ear of a 4-year-old male mon- key. Cell cycle was distributed by fluorescence-activated cell sorting when ceils were fixed and stained. As donor ceils, long-tailed macaque ear fibroblast cells were introduced into pig enucleated oocytes to investigate development of inter- species cloned embryos. [Result]Cell rate of G0/G1 phase under treatment tot 48 and 72 hours by 15 Ixmol/L Roscovitine （89.69%,90.49%） was significantly higher than the control group （P〈0.05）. The cell rate of G0/G1 phase was 91.12% and 90.46% when cells were treated by serum starvation and contact inhibition for 72 hours. Blastocyst formation rate was vast- ly improved under Roscovitine treatment for 48 hours, which was signifcantly higher than that treated by serum starvation and contact inhibition （P〈0.05）. [ Conclusion ]In conclusion, we suggested that roscovitine treatment of long-tailed macaque ear-derived fibroblastel cells can be used for synchronization of donor cells at the G1 or GO phase which improves the de- velopment of cloned monkey-porcine embryo.

关 键 词：猪 食蟹猴 异种核移植 细胞周期 同期化 胚胎发育 

分 类 号：Q813.6[生物学—生物工程]