不同电融合条件对小鼠卵丘细胞核移植重组胚融合和早期发育的影响

南方医科大学学报 ›› 2005, Vol. 25 ›› Issue (02): 156-159,164.

不同电融合条件对小鼠卵丘细胞核移植重组胚融合和早期发育的影响

江培洲^1,2, 沈新明^1,3, 乔贵林¹, 黄华¹, 姚开泰¹

1. 南方医科大学肿瘤研究所, 广东, 广州, 510515;
2. 南方医科大学组织学与胚胎学教研室, 广东, 广州, 510515;
3. 上海东方肝胆外科医院, 上海, 200438

出版日期:2005-02-20 发布日期:2005-02-20
基金资助:
收稿日期:2004-9-13。
基金项目:军队十五重点医学基金(98Z075);广东省自然科学基金(04300683)
作者简介:江培洲(1975- ),男,博士,讲师,电话:020-61648204,E-mail:jpzgmdxx@fimmu.com

Effects of different electrofusion parameters on activation and early development of mouse embryos reconstructed by cumulus cell nuclear transfer

JIANG Pei-zhou^1,2, SHEN Xin-ming^1,3, QIAO Gui-lin¹, HUANG Hua¹, YAO Kai-tai¹

1. 南方医科大学肿瘤研究所, 广东, 广州, 510515;
2. 南方医科大学组织学与胚胎学教研室, 广东, 广州, 510515;
3. 上海东方肝胆外科医院, 上海, 200438

Online:2005-02-20 Published:2005-02-20

摘要/Abstract

摘要： 目的研究不同电融合条件对来源于小鼠卵丘细胞核移植重组胚的融合和早期发育的影响,寻找最佳电融合参数。方法将直径10~12mm的C57BL/6j小鼠卵丘细胞核注射到去核卵母细胞透明带下,构建供体核-卵母细胞复合体。在不同电融合条件下诱导两者间的融合,并对重组胚激活以及随后发育的早期阶段包括2细胞、4~8细胞和桑椹胚进行观察和计数。结果电融合时电场强度或脉冲时间在一定范围内允许波动,变动范围分别是电场强度1000~2000kV/cm和脉冲时程40~160ms。低于容许范围的电融合参数会降低供体核-卵母细胞复合体的融合率,而高于容许范围的参数会导致供体核-卵母细胞复合体溶解乃至死亡。如果电融合参数在容许范围内,供体核-卵母细胞复合体的融合率无统计学差异,并且融合后重组胚的激活,以及发育至2细胞、4~8细胞和桑椹胚阶段的比率也无统计学差异。此外,脉冲重复次数以1~2次为佳。结论优化的电融合参数是决定供体核-卵母细胞复合体融合与重组胚激活的关键因素。

Abstract: Objective To study the effects of different parameters for electrofusion on the activation and early development of mouse embryos reconstructed by cumulus cell nuclear transfer, and explore optimal parameters for electrofusion. Methods A C57BL/6j mouse cumulus cell nucleus 10-12 mm in diameter was inserted into the perivitelline space of an enucleated oocyte. The fusion of donor-recipient pairs was induced with different parameters for electrofusion(with variation in electric field intensity, pulse duration and pulse times). Successful formation of the reconstructed embryos from donor-recipient pairs and the reconstructed embryos developing into the early embryonic stages(2-cell, 4-8-cell and morula stages) were observed and counted. Results The electric field intensity and pulse duration allowed variation during electrofusion within the range of 1 000-2 000 kV/cm and 40-160 ms, respectively. The donor-recipient pairs fused at very low rate when the parameters were below the allowed ranges, and disintegration or even death might occur when the parameters were above the ranges. Within these allowed ranges, variation of the electrofusion parameters did not produce significant impact on the ratio of the reconstructed embryos in 2-cell, 4-8-cell or morula stages(P<0.05). In addition, we suggested that pulse times be limited to 1-2. Conclusion Optimal parameters for electrofusion are crucial for the fusion of donor-recipient pairs and the activation of the reconstructed embryo.

中图分类号:

R329.25

江培洲^1,2, 沈新明^1,3, 乔贵林¹, 黄华¹, 姚开泰¹. 不同电融合条件对小鼠卵丘细胞核移植重组胚融合和早期发育的影响[J]. 南方医科大学学报, 2005, 25(02): 156-159,164.

JIANG Pei-zhou^1,2, SHEN Xin-ming^1,3, QIAO Gui-lin¹, HUANG Hua¹, YAO Kai-tai¹. Effects of different electrofusion parameters on activation and early development of mouse embryos reconstructed by cumulus cell nuclear transfer[J]. Journal of Southern Medical University, 2005, 25(02): 156-159,164.

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