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Cell-cycle synchronization of fibroblasts derived from transgenic cloned cattle ear skin: effects of serum starvation, roscovitine and contact inhibition

Published online by Cambridge University Press:  01 May 2008

XiuZhu Sun
State Key Laboratory for Agrobiotechnology, China Agricultural University, Beijing 100094, China.
ShuHui Wang
Institute of Animal Science Chinese Academy of Agricultural Science Beijing100094, China.
YunHai Zhang
AnHui Agricultural University, HeFei, 230036, AnHui province, China.
HaiPing Wang
Beijing GenProtein Biotechnology Ltd, BeiJing, 100094, China.
LiLi Wang
Beijing GenProtein Biotechnology Ltd, BeiJing, 100094, China.
Liu Ying
Beijing GenProtein Biotechnology Ltd, BeiJing, 100094, China.
Rong Li
Beijing GenProtein Biotechnology Ltd, BeiJing, 100094, China.
Ning Li*
State Key Laboratory for Agrobiotechnology, College of Biological Science China Agricultural University, Beijing 100094, China. State Key Laboratory for Agrobiotechnology, China Agricultural University, Beijing 100094, China.
All correspondence to: Ning Li. State Key Laboratory for Agrobiotechnology, College of Biological Science China Agricultural University, Beijing 100094, China. Tel: +86 10 62733323. Fax: +86 10 62733904. e-mail:


The purpose of the present study was to evaluate the effects of serum-starvation, contact-inhibition and roscovitine treatments on cell-cycle synchronization at the G0/G1 stage of ear skin fibroblasts isolated from transgenic cloned cattle. The developmental competence of re-cloned embryos was also examined. Our results showed that the proportion of G0/G1 cells from the serum-starved group at 3, 4 or 5 days was significantly higher compared with 1 or 2 days only (91.5, 91.7 and 93.5% versus 90.1 and 88.8%, respectively, p < 0.05); whilst there was no statistical difference among cells at 3, 4 or 5 days. For roscovitine-treated cells, the proportion of G0/G1 cells at 2, 3, 4 or 5 days was significantly higher than those treated for 1 day only (91.1, 90.1, 89.4 and 91.3% versus 86.51%, respectively, p < 0.05). The proportion of contact-inhibited G0/G1 cells rose significantly with treatment time, but was similar at 3, 4 and 5 days (89.4, 90.4, 91.4, 91.6 and 92.1%, respectively, p < 0.05). The efficiency of obtaining G0/G1 phase cells was lower when roscovitine treatment was employed to synchronize the cell cycle compared with the serum-starvation and contact-inhibition methods (89.7 versus 91.1% and 91.0%, p < 0.05). Moreover, obvious differences were observed in the rate of fused couplets and blastocysts (89.88 ± 2.70 versus 87.40 ± 5.13; 44.10 ± 8.62 versus 58.38 ± 13.28, respectively, p < 0.05), when nuclear transfer embryos were reconstructed using donors cells that had been serum starved or contact inhibited for 3 days. Our data indicate that 3 day treatment is feasible for harvesting sufficient G0/G1 cells to produce re-cloned transgenic bovine embryos, regardless of whether serum-starvation, contact-inhibition or roscovitine treatments are used as the synchronization methods.

Research Article
Copyright © Cambridge University Press 2008

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