聚合嵌合法制作黄牛和水牛种间嵌合胚的初步研究

A Preliminary Study on Making Interspecific Chimeras between Cattle and Buffalo by Aggregating Blastomeres

通过聚合嵌合法初步建立一套黄牛和水牛种间嵌合的程序与方法。聚合嵌合法采用链酶蛋白酶消化透明带或用机械剥离法去除透明带,然后在含有100μg/ml PHA的培养液中聚合形成嵌合胚。结果发现,8-细胞黄牛胚胎聚合水牛桑椹胚与黄牛囊胚聚合水牛桑椹胚相比,聚合胚存活率和囊胚发育率均无显著差异(P>0.05)。采用显微手术法分离黄牛和水牛8-细胞胚胎卵裂球进行聚合,聚合率为92.3%,囊胚发育率为58.3%,与用0.25%链酶蛋白酶分离胚胎卵裂球进行胚胎聚合的聚合率(86.7%)和囊胚发育率(46.2%)均无显著差异(P>0.05)。以上结果表明:①水牛和黄牛胚胎通过卵裂球聚合获得的种间嵌合胚胎能继续发育;②胚胎聚合前的发育阶段对其聚合成功率和随后的胚胎发育无明显影响。③胚胎卵裂球的分离方法(显微手术法和酶消化法)对其聚合率和囊胚发育率无明显影响。

To establish a preliminary procedure for making interspecific chimeras between cattle and buffalo by Aggregating Blastomeres Method. Chimeric embryos were reconstructed by aggregating blastomeres from early bovine and buffalo embryos isolated by either enzymatic or mechanical method in a droplet of culture medium containing 100 μg/ml phytohemagglutinin （PHA）. Aggregating buffalo morulae with bovine embryos at either 8-cell or blastula stage did not differ significantly in their aggregation rate （85.7% vs 77. 8%, P〉0. 05） and proportion developing to blastocysts （72.2% vs 42% P〉0. 05）. There were also no significant difference in aggregation rate （92.3% vs 86.7%, P〉0. 05） and blastocyst development （58.3% vs 46.2% P〉0.05） between the chimeric embryos derived from 8-cell embryonic blastomeres isolated by 0.25% Pronase and microsurgery. In conclusions,①The aggregation rate and embryonic development of chimeric embryos are not influenced by the stages of embryos to be aggregated;②Methods for isolation of blastomeres （enzymatic treament and microsurgery） do not affect their aggregation rate and embryonic development.