几种克服昆明小鼠 2- 细胞胚胎发育阻滞的培养液研究

SEVERAL MODIFIED CULTURE MEDIA CAN SUPPORT THE EMBRYOS OVERCOMING THE 2-CELL BLOCK AND DEVELOPING TO BLASTOCYST STAGE IN KUNMING MOUSE

比较了几种培养液对克服昆明小鼠胚胎 2 细胞发育阻滞的效果。实验 1的结果表明 ,在M16及CZB培养液基础上 ,加减几种成分得到的改进M16培养液 (用mM 16表示 )和改进的CZB培养液 (用mCZB表示 )均能有效克服 2 细胞阻滞现象。除去M 16和CZB培养液中的葡萄糖和磷酸盐后添加 5 5 5mmol/L (1g/L)果糖、2 5mmol/L牛磺酸、 10 0或110 μmol/LEDTA、 2mmol/L谷氨酰胺和 2 %必需氨基酸 (EAA)及 1%非必需氨基酸(NEA) ,均可支持体外培养的昆明小鼠 1 细胞胚胎发育到囊胚和孵化囊胚阶段。实验结果同时证明 ,TE培养液也具有良好的培养效果。 1 细胞胚胎在mM 16、TE和mCZB中培养后发育到囊胚的比率分别为 85 %、 85 %和 6 8%。实验 2的结果表明 ,在M 16培养液中单独添加 2 5mmol/L的牛磺酸也可支持体外培养的昆明小鼠 1 细胞胚胎发育到囊胚。在M16中同时添加 2 5mmol/L牛磺酸和 10 0 μmol/LEDTA两种成分可使 1 细胞胚胎的囊胚发育率达到 6 7%。通过比较分析我们认为牛磺酸对克服昆明小鼠胚胎 2 细胞阻滞起关键作用 。

The embryos of Kunming mouse, which is one of the most common experimental animals used in China, show cell block in vitro. The effects of several culture media on overcoming the 2cell block in embryo development were studied. In experiment 1, the modified M16 and CZB, in which the glucose and phosphate were excluded and 555mmol/L(1g/L) fructose, 100μmol/L or 110μmol/L EDTA, 25mmol/L taurine, 2mmol/L glutamine and 2% essential amino acid(EAA, 50×) and 1% nonessential amino acid(NEA, 100×) were added, were shown to overcome the 2cell block and support the development to blastocyst stage in 85% and 65% of the cultured 1cell embryos. The TE medium, in which EDTA, taurine and BSA were added, also supported the development to blastocysts in 85% of the 1cell embryos. In addition, the above three media could also support the development of 1 cell embryos to hatching blastocyst stage. In experiment 2, the medium of M16, in which 25mmol/L taurine or 25mmol/L taurine and 100μmol/L EDTA were added, was also shown to be effective to overcome the 2cell block, and 22% and 67% of cultured embryos developed to blastocyst stage respectively after 96 hours culture. We conclude that taurine plays an important role in overcoming the 2cell block during Kunming mouse embryo development.