CRBP-1基因RNAi慢病毒载体的构建与鉴定

Construction and identification of lentiviral vector of RNA interference of CRBP-1 gene

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 interfer-ence,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。

Objective ：To construct a lentivirul vector of RNA interference （RNAi） of rut cellular retinol-binding protein I （CRBP-I） gene. Methods ：The effective sequence of siRNA targeting CRBP-I gene was confirmed in our previous study. The complementary 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 lentivirul 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 successfully.