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目的:通过在非洲爪蛙胚胎中过表达ETS1探讨该基因对爪蛙胚胎早期发育的影响。方法:将克隆PCR获得的ETS1全长片段连接到质粒上,获得pCS2+-ETS1质粒;利用RT-PCR和Western blot检测ETS1 mRNA和蛋白在胚胎发育不同时期的表达情况;显微注射ETS1 mRNA入爪蛙胚胎,利用整胚原位杂交、定量PCR检测胚层标志性基因的表达;通过Western blot检测过表达ETS1对细胞凋亡和增殖的影响。结果:ETS1从受精卵时期即已开始表达,在器官形成期表达增强,过表达ETS1抑制了细胞分化以及神经胚期中胚层和外胚层标志基因表达,凋亡相关蛋白表达增加,增殖相关蛋白无明显变化。结论:ETS1表达于非洲爪蛙胚胎发育各阶段,过表达ETS1抑制了中胚层和外胚层发育,促进细胞凋亡但不影响细胞增殖。
OBJECTIVE: To investigate the effect of this gene on the early development of Clawed frog embryos by overexpression of ETS1 in Xenopus laevis embryos. Methods: The full-length ETS1 fragment obtained by cloning PCR was ligated into the plasmid to obtain pCS2 + -ETS1 plasmid. The expression of ETS1 mRNA and protein at different stages of embryo development was detected by RT-PCR and Western blot. The frog embryo was harvested for the detection of the germ layer by using in situ hybridization and quantitative PCR. The effect of overexpression of ETS1 on apoptosis and proliferation was detected by Western blot. Results: ETS1 began to express from the stage of fertilized egg and enhanced during organogenesis. ETS1 overexpression inhibited cell differentiation and the expression of germline and ectoderm marker genes in neuroblastoma, increased the expression of apoptosis related proteins and had no significant correlation with proliferation related proteins Variety. CONCLUSION: ETS1 is expressed in all stages of embryonic development of Xenopus laevis, overexpression of ETS1 inhibits mesoderm and ectoderm development, promotes apoptosis but does not affect cell proliferation.