摘要
目的为了研究X连锁凋亡抑制蛋白(X-linked inhibitor of apoptosis protein,XIAP)对前列腺癌的作用,构建靶向XIAP基因的小发夹RNA(small hairpin RNA,shRNA)表达载体.方法设计3对针对XIAP基因不同位点的shRNA片段,构建携带此shRNA片段的真核表达载体pGPU6/GFP/Neo-XIAP,分别命名为XIAP-shRNA1、XIAP-shRNA2和XIAP-shRNA3,进行酶切及测序鉴定.通过脂质体转染人前列腺癌细胞株DU145,并优化转染率.结果测序证明干扰序列完全正确,当质粒与脂质体比例1μg:2μl时转染率最高.结论成功构建重组质粒pGPU6/GFP/Neo-XIAP,为进一步研究XIAP基因在前列腺癌基因治疗奠定了基础.
Objective To investigate the effect of XIAP on prostate cancer,the XIAP-shRNA eukaryotic expression vectors were constructed.Methods Three pairs of shRNA that target the XIAP gene were designed.The eukaryotic expression vectors of pGPU6/GFP/Neo-XIAP named XIAP-shRNA1,XIAP-shRNA2,XIAP-shRNA3 were constructed and then were identified by restrictive digestion and DNA sequencing.Optimized transfection rate after transfected human prostate cancer cell line DU145 via liopsomes.Results Sequencing suggested that RNAi eukaryotic expression vectors targeting XIAP possessed correct nucleotide sequence.When the ratio of plasmid and liposomes is 1μg:2μl,transfection rate is the highest.Conclusions Recombinant pGPU6/GFP/Neo-XIAP shRNA plasmids are successfully constructed,which paves a way for continuous studying of XIAP function in gene therapy of prostate cancer.
作者
娄禄
徐觉剑
孙方浩
魏晋
李文智
Lou Lu;Xu Juejian;Sun Fanghao;Wei Jin;Li Wenzhi(Department of Urology,Xuzhou Muncipal Hospital Affiliated to Xuzhou Medical University,Xuzhou,Jiangsu,221002,China;Department of Urology,Shanghai Ninth People’s Hospital,Shanghai Jiao Tong University School of Medicine,Shanghai,200011,China)
出处
《泌尿外科杂志（电子版）》
2020年第3期46-51,共6页
Journal of Urology for Clinicians(Electronic Version)
关键词
X连锁凋亡抑制蛋白
RNA干扰
小发夹RNA
前列腺癌
X-linked inhibitor of apoptosis protein(XIAP)
RNA interference
Small hairpin RNA(shRNA)
Prostate cancer
作者简介
通信作者:娄禄,Email:loulu0325@163.com;通信作者:李文智,Email:drmrna@126.com
