盘羊供体细胞对异种核移植效率的影响

Effects of Argali (Ovis ammon) Donor Cells on Interspecies Nuclear Transfer

【目的】利用核移植技术生产异种重构胚,研究供体细胞对异种核移植效率的影响。【方法】以新疆盘羊和绵羊耳皮肤组织建立成纤维细胞系,进行细胞遗传学分析;以来自不同个体、不同代次、不同汇合度和不同保存方式的盘羊成纤维细胞为核供体,当地绵羊卵母细胞为受体,进行异种重构胚的构建。【结果】来自3只盘羊耳皮肤成纤维细胞的异种重构胚的卵裂率与囊胚率均无显著差异;采用10代以内的细胞作为核供体,细胞代次不会影响重构胚的发育率;生长到70%～80%汇合的供体细胞重构胚的囊胚率显著高于刚完全汇合组和汇合2～4d组;将供体细胞4℃保存2d,虽降低了卵裂率,但囊胚率没有受到显著影响;盘羊的染色体数目为2n=56,核型式为4(M)+50(T),XY(T,M);绵羊的染色体数目为2n=54,核型式为6(M)+46(T),XY(T,M)。【结论】利用本试验中的3只盘羊耳成纤维细胞,培养到10代以内、70%～80%汇合后用于构建异种重构胚,能得到较高的早期胚胎发育率。

[Objective] The aim of this study was to investigate the effect of passage number, confluency and storage method of argali （Ovis ammon） fibroblasts from different individuals on development of embryos. [Method] The embryos were reconstructed by interspecies nuclear transfer using local ovine oocyte cytoplasm. Fihroblasts were derived from skin of adult argali sheeps＇ ear in Xinjiang while they were analysized by chromosome karyotype. [Result] There was no difference in the cleavage rate and blastocyst rate among three the argali sheep. The passage number of fihroblast cells within 10 passages didn＇t affect the development rate of reconstructed embryos; The blastocyst rate, which was produced using 70%-80% confluent fibroblast cells, was higher than those using full-confluency cells on 0 d and 2-4 d. The cleavage rate of embryos reconstructed using donor cells stored for 2 days at 4℃ was lower than that using donor cells from control group （P〈0.05）, but no difference was found in the proportion of blastocysts in total embryos. Chromosome number of argali （Ovis ammon） and local sheep is 56 and 54, respcctively, and the karyotype formula is 4（M）＋50（T）, XY（T, M） and 6 （M）＋46（T）, XY（T, M）, respectively. [Conclusion] The embryos, which were reconstructed by interspecies nuclear transfer using 70%-80% confluent fibroblasts within 10 passages from three argali sheep in Xinjiang, had higher development competence.