摘要
To investigate the effects of different states of donor cells on the development of reconstructed sheep embryos, we designed five treatments of donor cells, including cell passage, cell size, serum starvation, colchicine treatment and gene transfection. Results are as follows: (Ⅰ) Compared with 16-18 passage cells, the morula/blastocyst rate of 5-7 passage cells as donor nuclei was significantly higher (17.3% vs. 4.9%, P<0.05), suggesting the advantage of short-time cultured cells in supporting the development of reconstructed embryos. (Ⅱ) The morula/blastocyst rate of reconstructed embryos derived from medium cells (15-25μm) as donor nuclei was higher than that from large cells (25-33μm) and small cells (8-15μm)( 20.0% vs. 8.0%, 9.7%), indicating that reconstructed embryos from medium cells had a greater potentiality to develop into morula/blastocysts than those from small or large ones. (Ⅲ) The morula/blastocyst rate of reconstructed embryos from donor cells of SS (serum starvation) was lower than that from donor cells of NSS (non-serum starvation), but no significant difference was detected between SS and NSS(11.8% vs. 18.6%, P>0.05). (Ⅳ) Fetal fibroblasts treated with 0.05μmol/L colchicine exhibited a higher morula/blastocyst rate of reconstructed embryos than those treated with 0.10 μmol/L colchicine and untreated ones (27.5% vs. 12.1%, 17.1%), however, no significant difference among the three treatments was detected (P>0.05). (Ⅴ) The morula/blastocyst rate of reconstructed embryos from fetal fibroblasts transfected with GFP gene only was 3.1%, significantly lower than that from non-transgenic cells (3.1% vs. 20.4%, P<0.05). In conclusion, our results demonstrated that fetal fibroblasts of fewer passages, medium size could ensure a higher morula/blastocyst rate of reconstructed embryos. Serum starvation of donor cells might be unnecessary to the development of reconstructed embryos. Donor cells treated with 0.05μmol/L colchicine could facilitate the development of reconstructed embryos. Additionally, as cells transfected with GFP gene were used as donor nuclei, adverse effect on the development of reconstructed embryos was observed. Therefore, the developmental efficiency of reconstructed embryos could be improved if proper treatments to donor cells were used.
To investigate the effects of different states of donor cells on the development of reconstructed sheep embryos, we designed five treatments of donor cells, including cell passage, cell size, serum starvation, colchicine treatment and gene transfection. Results are as follows: (Ⅰ) Compared with 16-18 passage cells, the morula/blastocyst rate of 5-7 passage cells as donor nuclei was significantly higher (17.3% vs. 4.9%, P<0.05), suggesting the advantage of short-time cultured cells in supporting the development of reconstructed embryos. (Ⅱ) The morula/blastocyst rate of reconstructed embryos derived from medium cells (15-25μm) as donor nuclei was higher than that from large cells (25-33μm) and small cells (8-15μm)( 20.0% vs. 8.0%, 9.7%), indicating that reconstructed embryos from medium cells had a greater potentiality to develop into morula/blastocysts than those from small or large ones. (Ⅲ) The morula/blastocyst rate of reconstructed embryos from donor cells of SS (serum starvation) was lower than that from donor cells of NSS (non-serum starvation), but no significant difference was detected between SS and NSS(11.8% vs. 18.6%, P>0.05). (Ⅳ) Fetal fibroblasts treated with 0.05μmol/L colchicine exhibited a higher morula/blastocyst rate of reconstructed embryos than those treated with 0.10 μmol/L colchicine and untreated ones (27.5% vs. 12.1%, 17.1%), however, no significant difference among the three treatments was detected (P>0.05). (Ⅴ) The morula/blastocyst rate of reconstructed embryos from fetal fibroblasts transfected with GFP gene only was 3.1%, significantly lower than that from non-transgenic cells (3.1% vs. 20.4%, P<0.05). In conclusion, our results demonstrated that fetal fibroblasts of fewer passages, medium size could ensure a higher morula/blastocyst rate of reconstructed embryos. Serum starvation of donor cells might be unnecessary to the development of reconstructed embryos. Donor cells treated with 0.05μmol/L colchicine could facilitate the development of reconstructed embryos. Additionally, as cells transfected with GFP gene were used as donor nuclei, adverse effect on the development of reconstructed embryos was observed. Therefore, the developmental efficiency of reconstructed embryos could be improved if proper treatments to donor cells were used.
作者
WANG Hai, AO Hong, PAN QiuZhen, LI RongQi, ZHAO MengBin, LI AN ZhengXing, LI Ning & WU ChangXin 11nstitute of Zoology, Chinese Academy of Sciences, Beijing 100080, China
2 Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100094, China
3 Northeast Agricultural University, Harbin 150030, China
4Beijing Glorious Land Agricultural Co., Ltd. Beijing 100049, China
5 College of Animal Science and Technology, China Agricultural University, Beijing 100094, China
6 State Key Laboratory of Agrobiotechnology, China Agricultural University, Beijing 100094, China
基金
the National Key Basic Research '973' Program of China (Grant No. 2006CB102I00)
