Stella基因真核表达载体的构建及其对小鼠胚胎干细胞多能性的影响
Construction of Stella Eukaryotic Expression Vector and Its Effect on Pluripotency of Mouse Embryonic Stem Cells
白耀富1#，刘维帅2#，华进联1*
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作者单位：(1. 西北农林科技大学动物医学院，陕西省干细胞工程技术中心，农业部动物生物技术重点实验室，陕西杨凌 712100；2. 杨棱示范区医院病理科，陕西杨凌 712100)
中文关键字：胚胎干细胞；减数分裂；多能性
英文关键字：embryonic stem cells; meiosis; pluripotency
中文摘要：构建Stella基因真核表达质粒，转染小鼠胚胎干细胞(Embryonic stem cells，ESC)并初步探讨Stella对减数分裂起始相关基因(Stra8)及胚胎干细胞多能性的影响。通过RT-PCR扩增目的基因，并连接至真核表达载体pEGFP-C1，利用重组质粒转染小鼠胚胎干细胞。对转染细胞进行荧光检测，确认Stella的表达，并利用免疫荧光及PCR检测转染细胞基因表达情况。酶切鉴定及测序分析表明成功构建含Stella基因的重组真核表达质粒，过表达Stella对ES细胞的增殖和形态学特征、进入减数分裂阶段的相关基因及其多能性基因的表达影响并不显著。故此得出结论：Stella在小鼠胚胎干细胞中能够正确表达，但对ES细胞的分化、Stra8基因的表达及其多能性基因的表达并无显著影响。
英文摘要：To explore the effects of recombinant vector Stella on the pluripotency of mouse embryonic stem cells (ESCs) and expression profile of specific meiosis marker-Stra8 in transfected ESCs. The eukaryotic expression vector containing Stella was constructed and transfected into ESCs. Expression profiles of GFP was observed in transfected ESCs, and expression levels of pluripotent genes and specific meiosis markers were simultaneously detected by RT-PCR and immunofluorescent staining. The results demonstrated that although Stella which had been cloned in the recombinant vector was successfully expressed in ESCs as identified by GFP and PCR amplification, it had no significant effect on the expression profiles of pluripotent and specific meiosis marker genes.