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| Investigation on Affects of Different Supplements in the Medium on Isolation and Culture of Embryonic Stem Cells from Mouse Embryos of Kunming Species |
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Abstract This experiment focued on investigating affects of different supplements in the media containing KSR, FBS, FBS+PD98059, respectively on embryo attachment, formation of ICM outgrowths, isolation and culture of embryonic stem cells (ESCs) from mouse embryos of Kunming species, which was used as the experimental material while mitomycin C-inactivated mouse embryonic fibroblasts (MEF) as the feeder layers. The results demonstrated that the efficiencies of embryo attachment in the medium supplied with KSR was significantly lower compared with that of the medium supplied with FBS（P＜0.05), and formation efficiencies of ICM outgrowths and ESCs clonies at the first passage appeared no significant difference between them (P>0.05), however, formation efficiencies of ESCs clonies at the 2~5 passages were significantly higher than those in the medium supplemented with FBS (P＜0.05), two ESCs lines were subcultured to the seventh passages. Meanwhile, the efficiencies of embryo attachment, formation of ICM outgrowths and ESCs clonies at the 1-5 passages in the medium supplemented with FBS + PD98058, were significantly lower than those in the medium supplemented with KSR and FBS, respectively（P＜0.05). Providing ICM cells and ESCs were trypsinized with 0.5g/L trypsin +0.2g/L EDTA combined with manual dissection, ESCs clonies at the 1~5 passages appeared significantly higher than those with 2.5g/L trypsin + 0.2g/L EDTA （P＜0.05). Therefore, the results indicated that it was advantageous to isolation and culture of ESCs from mouse embryos of Kunming Species in the medium supplemented with KSR over FBS and FBS+PD98059, respectively. Accordingly, the ICM cells and ESCs at the initial passages were trypsinized more properly with 0.5 g/L trypsin associated with manual dissociation, than with 2.5 g/L trypsin.
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Received: 02 May 2007
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