12/24/2023 0 Comments Siberian mouse analIn generally, Xenopus laevis oocytes at the germinal vesicle (GV) stage are extremely larger than mammalian oocytes and accessible with relative ease. Due to the limitation in accessibility of mammalian oocytes, non-mammalian eggs the large number and volume would be a tempting alternative. Furthermore, these cells showed up-regulation in the expression of pluripotency markers ( Oct4, Sox2, and Nanog) and formed outgrowth colonies which similar with morphology of embryonic stem cells (ESCs) ( Miyamoto et al., 2008).Īs mentioned above, the use of totipotent or pluripotent cell extract for pre-treatment of donor cells prior to SCNT has been expected to improve epigenetic reprogramming of cloned embryos and eventually enhance the frequency of development to term. It has been reported that the transcriptional reprogramming of human and bovine nuclei increased after treatment of cells in extracts from Xenopus laevis oocytes or egg ( Hansis et al., 2004 Alberio et al., 2005). (2007) has been reported that porcine metaphase (MII) oocyte extract replaces transcription factors from donor nuclei with the oocyte extract and eventually increases the histone deacetylation in the somatic nuclei. When ovine SCNT embryos reconstructed by using donor cells pretreated with Xenopus laevis germinal vesicle (GV) oocyte extracts were transferred into surrogate, the pregnancy and survival rate were greatly improved ( Rathbone et al., 2010). In the pioneering studies with amphibians and mammals, it was demonstrated that epigenetic reprogramming of differentiated mammalian cells were successfully induced to a pluripotent state by exposing amphibian oocyte extracts ( Hochedilinger et al., 2002 Alberio et al., 2005 Bian et al., 2009). The ex-ovo system for epigenetic reprogramming of a terminately differentiated cell depends on the transient uptake of regulatory components from a nuclear and cytoplasmic mixtures derived from cell extract ( Håkelien et al., 2002 Landsverk et al., 2002). Keywords: Epigenetic Modifications Somatic Cell Nuclear Transfer Siberian Sturgeon Oocyte Extract Our results suggested that pre-treatment of cells with SOE could improve epigenetic reprogramming and the quality of porcine SCNT embryos. And there was observed a better development to the blastocyst stage in the SOE-treated group (p<0.05). The number of apoptotic cells was significantly decreased and pluripotency markers ( Nanog, Oct4 and Sox2) were highly expressed in the blastocyst stage of SCNT embryos reconstructed using SOE-treated cells as nuclear donor (p<0.05). Additionally, the level of histone acetylation in SCNT embryos at the zygote stage was significantly increased when reconstructed using SOE-treated cells (p<0.05), similar to that of IVF embryos at the zygote stage. As results, the intensity of H3K9ac staining in PFFs following treatment of SOE for 7 h at 18☌ was significantly increased but the intensity of H3K9me3 staining in PFFs was significantly decreased as compared with the control (p<0.05). A reversible permeability protocol with 4 μg/mL of digitonin for 2 min at 4☌ in order to deliver Siberian sturgeon oocyte extract (SOE) to porcine fetal fibroblasts (PFFs) was carried out. Here we hypothesized that treatment of somatic cells with extracts prepared from the germinal vesicle (GV) stage Siberian sturgeon oocytes prior to their use as nuclear donor for SCNT would improve in vitro development. However, because the efficiency of SCNT still remains low, a combination of SCNT technique with the ex-ovo method may improve the normal development of SCNT embryos. In the SCNT experiment, nuclear reprogramming is induced by exposure of introduced donor nuclei to the recipient cytoplasm of matured oocytes. Somatic cell nuclear transfer (SCNT) has generally demonstrated that a differentiated cell can convert into a undifferentiated or pluripotent state.
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