Heterogeneous Embryo Reconstruction In Camel Skin Fibroblast Cell

The research reconstructed three kinds of embryo, they were sheep-sheep,camel-sheep and camel-bovine. At first, camel and sheep skin fibroblasts were cultured in different ways as dorner cells and then, prepared the enucleated sheep and bovine oocytes to reconstruct embryos. So, research findings had been received by culturing embryos in vitro. Camel skin fibroblasts were dissociated and cultured in vitro. The primary cells were obtained by explant tissue culture at 37℃,100% humidity and 5% CO2, DMEM-F12 supplemented with 10% NCS were used to support the fibroblast growth. Fibroblasts culture had been done in 3 ways and also received the best one in this research. Trypsin digestion was feasible for separating homogeneous fibroblasts from the mixture of fibroblasts and epithial cells after repeatitial attachments for two or three times of subcultures. Trypsin dissolved in D-Hanks was suitable for digestion. The results of experiment showed at first the parthenogenetic activation of oocytes. Matured bovine and sheep oocytes were efficiently activated by 5mM Ionomycin together with 6-DMAP, and then the actived oocytes were cultured in SOFaa added with 10% FBS and cumuluses, a higher rate of blastocysts development were achieved (bovine 27.34%,sheep 46.08%). Under the condition of 20 μ second duration, 2 pulses and 0.5 seconds internal, the actived oocytes were cultured in SOFaa added with 10% FBS and cumuluses, when the voltage was near the 1.4KV/cm and the rates of blastocysts development reached peak values in bovine oocyte. But in sheep oocyte, the highest voltage was near about 1.5KV/cm. Certainly, bovine parthenogenetic activated blastocyst could be received when the amplitude was changed from 1.0 KV/cm to 2.4KV/cm and also sheep parthenogenetic activated blastocyst from 1.0 KV/cm to 2.2 KV/cm. The research indicated that the cloned embryos reconstructed by sub zona pellucida injection associated with electrofusion. Sheep-sheep reconstructed embryos were developed to blastocysts, camel-sheep reconstructed embryos to morula and camel-bovine reconstructed embryos to 8-16 cells stage.