文档介绍：CRBP-1基因RNAi慢病毒载体的构建与鉴定
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作者:陈波,李建平,戴途,吴志勇,罗蒙,孙勇伟
【摘要】目的:构建大鼠视黄醇结合蛋白-1(cellular retinol-binding protein- I ,CRBP-1)基因RNAi(RNA interference, RNAi)慢病毒载体。方法:针对已经筛选确定的大鼠CRBP-1 基因RNAi有效靶序列,合成靶序列的Oligo DNA,退火形成双链DNA,与经Hpa I 和Xho I酶切后的pGCL-GFP载体连接产生短发卡RNA慢病毒载体,PCR 筛选阳性克隆,测序鉴定。结果:PCR 鉴定与DNA测序证实合成的含CRBP-1 shRNA慢病毒载体寡核苷酸链插入正确。结论:成功构建大鼠CRBP-1基因RNAi 慢病毒载体。
【关键词】 RNA干扰;视黄醇结合蛋白-1;慢病毒
[Abstract] Objective:To construct a lentiviral vector of RNA interference(RNAi) of rat cellular retinol-binding protein I (CRBP-I) gene. Methods:The effective sequence of siRNA targeting CRBP-I gene was confirmed in our previous study. plementary DNA containing both sense and antisense Oligo DNA of the targeting sequence was designed,synthesized and cloned into the pGCL-GFP vector,to construct a lentiviral vector which expressed short hairpin RNA(shRNA), and it was identified by PCR and DNA sequencing. Results:PCR identification and DNA sequencing demonstrated that insertion of oligonucleotide of the lentivirus RNAi vector containing CRBP-I shRNA was right. Conclusion:The lentivirus RNAi vector of rat CRBP-I was constructed essfully.
[Key words] RNA interference;Cellular retinol-binding protein I;Lentivirus
肝星状细胞(hepatic stellate cells, HSC)的激活过程是肝脏胶原产生和肝纤维化形成的中心环节[1],在肝纤维化逆转过程中也起重要作用,2002年Uchio K[2]l4制备大鼠肝纤维化模型中发现,活化第5天的HSC中CRBP-1表达明显增加,而视黄醇脂滴消失,提示CRBP-1与肌