【摘要】 目的 提高小鼠体细胞核移植过程中融合效率。 方法 使用CRY-3型和CE-450型细胞融合仪对 B₆D₂F₁小鼠去核卵母细胞与该品系小鼠胎儿成纤维细胞构成的重构胚进行融合，通过PEG/DMSO 对供核细胞的预处理，比较了不同融合液，不同电极形状融合槽和供核细胞代次对融合率的影响。结果 （1） 以PEG/DMSO对供核细胞预处理5 min的融合率显著高于同条件下未处理组（51.11% vs 25.64%）（P<0.05）; （2）融合液中含0.1 g/L聚乙烯醇（PVA）组的融合率高于无PVA组; 重构胚在甘露醇浓度为0.32 mol/L融合液中的融合率（48.21%）高于其他甘露醇浓度组（27.78%,41.67%和30.11%）; （3） 使用针状电极融合槽时的融合率（42.31%, 43.13%）高于同条件下平行丝状电极融合槽组（30.43%, 33.33%）, 其中又以在2 000 V/cm, 20 μs, 2个脉冲时, 交流电预处理30 s组最高（43.13%）; （4）供核细胞传代一次后融合率随着传代次数的增加而下降。结论 通过PEG/DMSO对供核细胞进行预处理,并在融合液中添加 PVA 时，结合优化的融合条件可以提高小鼠体细胞核移植过程中融合率。更多还原

【Abstract】 Objective To improve the efficiency of electrofusion of reconstructed embryos derived from oocytes and fetal fibroblast cells both of B₆D₂F₁ strain within a mouse somatic cell nuclear transfer（SCNT） program. Methods The effect of different pre-treatment on donor cells,fusion midia, electrical field intensity, electrodes’ shape and cell passage on the efficiency of electrofusion was studied. Results（1）The pre-treatment of PEG/DMSO on donor cells significantly improved the fusion rate（51.11% vs 25.64%）（P<0.05）;（2） Fusion rate in the fusion media containing 0.1g/L polyvinyl alcohol（PVA） was higher than those in the media without PVA. Fusion rate in fusion media containing 0.32 mol/L mannitol（48.21%） was higher than those in the media with 0.28 mol/L, 0.30 mol/L and 0.36 mol/L mannitol（27.78%, 41.67% and 30.11%）;（3）Although the chamber is a much more routine way for fusing the reconstructed embryos, the tip shape electrodes can improve the fusion rate（42.31%, 43.13%） comparing to routine parallel line electrodes（30.43%, 33.33%）;（4） the efficiency of electrofusion of oocytes with fetal fibroblast cells at the found passage and the first passage was higher than that at the second and the third passage（38.98%, 38.46%, 36.73% and 35.00%）, regardless of other fusion conditions.Conclusion Both the pre-treatment of PEG/DMSO on donor cells and the addition of PVA to the fusion media can improve the efficiency of electrofusion in the procedure of mouse SCNT.更多还原