Department of Research in Animal Production, Laboratory of Animal Reproduction and Biotechnology, 07040 Olmedo, Sardinia, Italy.
Cell Biol Int. 2009 Dec 16;34(1):53-60. doi: 10.1042/CBI20090008.
Transcription factor Oct4 (octamer-binding transcription factor-4) is important in early embryonic development and differentiation. It is also required for maintenance of pluripotency of the inner cell mass, and is used as a staminality marker of embryonic stem cells. Changes in Oct4 expression during the different stages of early embryo development have been reported, and therefore we have conducted a quantitative study of Oct4 gene expression of sheep blastocysts in vitro, and of embryonic-stem-like cells at the undifferentiated stage and in the course of differentiation. To characterize embryonic-stem-like cells, alkaline phosphatase activity, stage-specific embryonic surface antigens SSEA-1, SSEA-3, SSEA-4 and three specific gene markers Nanog, Sox2 and Stat3 were assayed. cDNA produced by RT (reverse transcriptase)-PCR was synthesized and amplified by PCR; sequencing gave 98, 95 and 98% homology with the bovine sequences of Oct4, Nanog and Stat3 respectively. Using the ovine sequence of 290 bp, quantitative expression of Oct4 in the inner cell mass, trophoblast and embryonic-stem-like cells was performed by qRT-PCR (quantitative real-time PCR). Oct4 was expressed in the inner cell mass, trophoblast and embryonic-stem-like cells. Expression in the inner cell mass was significantly higher than in the trophoblast. This could be useful in defining the quality of embryos produced and makes it possible to use Oct4 to detect pluripotency. In addition, the different levels of Oct4 expression between undifferentiated and differentiating embryonic-stem-like cell cultures could be used to detect this gene as a staminality marker.
转录因子 Oct4（八聚体结合转录因子 4）在早期胚胎发育和分化中起着重要作用。它也是维持内细胞团多能性所必需的，并且被用作胚胎干细胞的干细胞标记物。已经报道了 Oct4 表达在早期胚胎发育的不同阶段的变化，因此我们对绵羊体外囊胚和未分化阶段以及分化过程中的胚胎样干细胞中的 Oct4 基因表达进行了定量研究。为了表征胚胎样干细胞，检测了碱性磷酸酶活性、胚胎特异性表面抗原 SSEA-1、SSEA-3、SSEA-4 和三个特异性基因标记物 Nanog、Sox2 和 Stat3。通过 RT（逆转录酶）-PCR 合成并扩增了 cDNA；测序与牛 Oct4、Nanog 和 Stat3 序列的同源性分别为 98%、95%和 98%。使用 290 bp 的绵羊序列，通过 qRT-PCR（定量实时 PCR）对 Oct4 在内细胞团、滋养层和胚胎样干细胞中的定量表达进行了研究。Oct4 在内细胞团、滋养层和胚胎样干细胞中表达。内细胞团中的表达显著高于滋养层。这对于定义所产生胚胎的质量很有用，并使得可以使用 Oct4 来检测多能性。此外，未分化和分化的胚胎样干细胞培养物之间 Oct4 表达水平的差异可用于检测该基因作为干细胞标记物。
