细胞因子信号抑制蛋白-3基因RNA干扰慢病毒载体的构建

发布时间：2018-04-16 23:35

  本文选题：细胞因子信号抑制蛋白3 + RNA干扰　； 参考：《吉林大学》2012年硕士论文

【摘要】：支气管哮喘是以Th2细胞增多占优势，同时伴随肥大细胞、嗜酸性粒细胞联合浸润为特征的一种慢性疾病，Th1/Th2的比例失衡和以Th2型细胞为主的免疫应答在支气管哮喘的发病机制中起到重要作用。SOCS-3是JAK/STAT信号通路的反馈抑制因子，又被称为细胞的“分子刹车”。SOCS-3与多种疾病的发生发展有着密切关系，同时亦是多条信号通路的交界点，从而其调控、作用机制复杂。SOCS-3不仅是细胞因子的调节者，而且在调控细胞分化和决定细胞分化方面起着重要作用。SOCS-3在Th2细胞中高表达，并可被许多炎症或抗炎症因子诱导生成，，同时能反作用于Th细胞，影响其分化。 本实验根据小鼠SOCS3的mRNA序列设计3条shRNA序列，化学合成后与PLL3.7质粒连接，产生PLL3.7-SOCS-3-shRNA慢病毒重组质粒，经体外转染小鼠B16黑色素瘤细胞鉴定其沉默效率，选取沉默效率最高者与慢病毒包装质粒共转染293T细胞，经收集细胞上清和低温超高速离心，最终获得高浓度的慢病毒载体。本实验构建SOCS-3RNA干扰慢病毒载体，以期为后续的支气管哮喘的研究与治疗提供实验工具。
[Abstract]:Bronchial asthma is dominated by the proliferation of Th2 cells, accompanied by mast cells.The imbalance of Th1 / Th2 ratio in a chronic disease characterized by eosinophilic granulocyte infiltration and the immune response with Th2 type cells play an important role in the pathogenesis of bronchial asthma. SOCS-3 is a feedback inhibitor of JAK/STAT signaling pathway.SOCS-3, also known as cell "molecular brake", is closely related to the occurrence and development of many diseases, and it is also the junction point of many signal pathways. Therefore, the complex mechanism of SOCS-3 is not only the modulator of cytokines, but also the junction of many signal pathways.SOCS-3 plays an important role in regulating cell differentiation and determining cell differentiation. SOCS-3 is highly expressed in Th2 cells and can be induced by many inflammatory or anti-inflammatory factors.In this study, three shRNA sequences were designed according to the mRNA sequence of mouse SOCS3, which were chemically synthesized and ligated with PLL3.7 plasmid to produce PLL3.7-SOCS-3-shRNA lentivirus recombinant plasmid. The recombinant plasmid was transfected into murine B16 melanoma cells in vitro to identify its silencing efficiency.The highest silencing efficiency and lentivirus packaging plasmid were cotransfected into 293T cells. After supernatant and cryogenic centrifugation, a high concentration of lentivirus vector was obtained.In this study, SOCS-3RNA interference lentivirus vector was constructed to provide experimental tools for the further study and treatment of bronchial asthma.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R562.25

【参考文献】

中国期刊全文数据库 前2条

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2 祖莹;李成荣;李德发;邓继岿;郑跃杰;;细胞因子信号抑制因子在支气管哮喘T_H1／T_H2细胞失衡中的作用[J];中华微生物学和免疫学杂志;2006年06期

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