黄牛-奶牛IVF胚胎早期发育影响因素的研究

为了在实验室建立一套黄牛-奶牛IVF胚胎制备体系,为IVF胚胎的研究提供来源方便、制备简单的素材,本实验从黄牛-奶牛IVF胚胎制备过程中卵巢的处理方法、精子的处理方法及受精卵的培养方法等几个方面进行优化。结果表明：温度递增式洗涤卵巢(25℃、30℃、35℃、39℃),对卵母细胞起到一定的缓冲作用,获取A级COC_S的比例和卵母细胞的成熟率较直接用25℃和39℃生理盐水洗涤卵巢高且差异显著,但是卵裂率和囊胚率并没有显著差异;用移液器轻轻吹打精子后,显著提高卵母细胞的受精率和卵裂率;培养方法对黄牛-奶牛IVF胚胎的早期发育有一定的影响。研究结果提示,经过温度递增式的生理盐水梯度洗涤卵巢,以体外成熟的黄牛卵母细胞为受体、以高产奶牛冷冻精子经过移液器轻微反复吹打处理后进行体外受精,受精卵置五孔板中进行IVF胚胎的体外培养,可以获得发育至孵化囊胚的黄牛-奶牛IVF胚胎。

Influencing Factors of Early Embryos Development of Cattle-cow in Vitro Fertilization

To establish a set of cattle-cow IVF embryo preparing system in laboratory and provide convenient sources and simple materials for IVF embryo research, several aspects such as ovarian treatment, sperm treatment and the culture conditions of fertilized eggs were optimized. The results showed that ovaries washed with increasing temperature (25℃, 30℃, 35℃, 39℃) had buffer effects on the oocytes, and higher proportion of A-COCs and matured oocytes were obtained compared with that of washing ovaries directly with 25℃ and 39℃ normal saline, and the difference was significant, but there was no significant difference with the cleavage rate and blastocyst rate. After blowing sperm gently with pipette, the fertilized oocytes and cleavage rate were significantly increased. Culture method had a certain effect on the early development of IVF embryo. The results showed that the hatching blastocysts could be obtained by in vitro fertilization of the mature cattle oocytes and the frozen spermatozoa of the high-yield dairy cows, which are treated with the pipette and blown repeatedly and the fertilized eggs are placed in a five-hole plate for IVF embryo culture in vitro.