摘要:
ERK5通路是丝裂原活化蛋白激酶(MAPK)信号通路家族成员之一,是一种非典型的MAPK通路,也叫做大MAPK通路(Big MAPK/BMK1),可以被各种刺激因素激活,与多种细胞反应(如细胞增殖、分化、转化及凋亡等)及机体多种生理病理过程(如脑发育、癌症及糖尿病等)密切相关.采用RNA干扰的方法,用构建的ERK5干扰质粒干扰ERK5表达,通过RT-PCR和Western-blotting的方法分别在RNA和蛋白水平上来检测ERK5的表达,鉴定干扰效率;通过G418对转染了干扰质粒的鼠恶性黑色素瘤细胞(B16F10细胞)进行稳定筛选,得到ERK5 RNA干扰的B16F10稳定细胞系,为后期的动物实验做准备.
Abstract:
ERK5 (extracellular-regulated kinase 5) signal pathway is a member of the MAPK family. It is an atypical MAPK signal pathway and is also named Big MAPK/BMK1. ERK5 signal pathway can be activated by various stimulating factors, and is closely associated with many kinds of cell responses, such as proliferation, differentiation, transformation and apoptosis. It is also related to the processes of physiological and pathological reactions including the development of brain, cancer and diabetes. In this study,RNAi (RNA interference) was applied to decrease the expression of ERK5. The expression of ERK5 was detected by RT-PCR and Western-blotting. B16F10 cells transformed by constructed ERK5 shRNA plasmids were stably screened using G418, and an ERK5 RNAi B16F10 stable cell line was obtained, whichwas ready for further animal experiments.
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