中国农业科学 ›› 2007, Vol. 40 ›› Issue (11): 2607-2614 .

• 畜牧·兽医·资源昆虫 • 上一篇    下一篇

水牛腔前卵泡体外培养效果影响因素的研究

冯贵雪,杨素芳,潘红平,卞桂华,石德顺   

  1. 广西大学动物繁殖研究所
  • 收稿日期:2006-06-23 修回日期:2006-10-24 出版日期:2007-11-10 发布日期:2007-11-10
  • 通讯作者: 石德顺

A study on the in vitro culture of buffalo preantral follicles

  

  1. 广西大学动物繁殖研究所
  • Received:2006-06-23 Revised:2006-10-24 Online:2007-11-10 Published:2007-11-10

摘要: 【目的】主要探讨水牛腔前卵泡体外培养的影响因素。【方法】采用梳刮法从沼泽型水牛的卵巢中回收得到腔前卵泡,而后用McCoy's5a培养液在96孔培养板中体外培养5~10 d,在显微镜下观测腔前卵泡的形态与增长情况,并用台盼蓝染色鉴定腔前卵泡的存活情况。【结果】在培养液中添加100 µg•L-1的FSH能显著提高腔前卵泡培养5和10 d后的增长幅度,当同时添加50 mg•L-1的维生素C时增长幅度进一步加强;添加50 mg•L-1的维生素C能提高腔前卵泡培养10 d后的形态正常率,但对卵泡增长及存活没有影响;添加50 µg•L-1的EGF能提高腔前卵泡培养5 d的增长幅度,但对培养10 d后腔前卵泡的作用不显著。随着卵泡的直径增加,体外存活率升高,增长的幅度上升,异常率下降。60~100 µm的腔前卵泡用三维培养法(琼脂糖包埋)体外培养10 d的直径增长幅度显著高于二维培养法(琼脂糖铺底)和普通培养法,且形态异常显著低于二维培养法和普通培养法;100~140 µm卵泡用二维培养法培养的增长幅度最大,培养成活率最高。【结论】FSH能促进水牛卵泡的体外增长,维生素C能维持卵泡的正常形态结构,且两者的协同作用显著;水牛卵泡体外发育能力随其体积的增大而增强;三维培养法适宜于培养60~100 µm的卵泡,而二维培养法有利于100~140 µm卵泡的体外发育。

关键词: 水牛, 腔前卵泡, 体外培养

Abstract: Growth of buffalo preantral follicles in vitro culture was studied to investigate the effects of size of preantral follicles, methods of culture, relative factors. In experiment 1, Effects of ascorbic acid (Vc), FSH and EGF on the in vitro development of buffalo preantral follicles were examined. Ascorbic acid can maintain the follicular morphologic integrity during culture but does not affect the survival and growth of follicles. FSH can promote the growth of follicles, but does not affect the survival and morphology of follicles. The growth promotive role of FSH is enhanced by the presence of Vc. EGF can accelerate the diameter increasing of follicles in the early culture stage, and this action is enhanced in the presence of FSH. In experiment 2,The in vitro growth of follicles in different diameters (60~180µm) was examined. As the follicular diameter increases, the survival rate and growing extent of follicles increases, the morphological abnormity of follicles decreases. In experiment 2, Conventional, two dimension (agarose coating) and three dimension (agarose embed) culture system were compared in their efficiency of supporting follicle growth. Three dimensions culture system is proved to be suitable for culturing the follicles in diameter of 60~100µm, while the two dimension culture system is proved to be suitable for culturing the follicles in diameter of 100~140µm.

Key words: buffalo , preantral follicle, in vitro culture