针对MAGE-1的pSUPERRNAi系统的构建  

Construction of pSUPER RNAi system of inhibiting MAGE-1 activity.

作　　者：程光[1] 章翔[1] 张赟[2] 鲍炜[2] 曹卫东[1] 高大宽[1] 宋蕾[1] 

机构地区：[1]第四军医大学西京医院全军神经外科研究所,陕西西安710032 [2]第四军医大学西京医院免疫学教研室,陕西西安710032

出　　处：《现代肿瘤医学》2006年第3期261-264,共4页Journal of Modern Oncology

基　　金：国家自然科学基金资助项目(编号39970752)

摘　　要：目的构建抑制MAGE-1的siRNA表达载体。方法化学合成2对编码短发夹RNA序列的、靶向MAGE-1基因的寡核苷酸链,各64个碱基,退火,克隆到经BglⅠ、HindⅢ双酶切的pSUPER载体上,重组构建RNAi质粒。通过双酶切鉴定及基因片段序列分析验证构建效果。结果重组构建的pSUPER-MAGE-1载体经双酶切电泳分析及插入基因片段序列分析,结果表明64个碱基成功插入到预计位点,并且序列完全一致。结论载体的成功构建,为进一步研究MAGE-1在肿瘤中的功能打下基础,可用于分析基因功能、肿瘤治疗等。Objective To construct expressing vector of siRNA in order to inhibit MAGE - 1 activity. Methods Sixty four base - pair oligos for hairpin RNA expression which targeted MAGE - 1 gene were chemically synthesized and annealed, pSUPER vector was linearized with Bgl Ⅱ and Hind Ⅲ. Finally, annealed oligos were inserted into the treated pSUPER to construct RNAi plasmid（pSUPER - MAGE - 1 ）. The reconstructed RNAi plasmid must be identiffed by electrophoresis after digestion with EcoR Ⅰ and Hind Ⅲ and be confirmed by sequencing analysis. Results Recombinant pSUPER - MAGE - 1 vector was identified by digestion with EcoR Ⅰ and Hind Ⅲ, and confirmed by sequencing analysis. The results demonstrated that 64 bp had been inserted the expected site. Furthermore, the insertion sequence was exactly correct. Conclusion pSUPER - MAGE - 1 RNAi system has been constructed successfully. This will facilitate the study of MAGE - 1 activity inhibition and function.

关 键 词：PSUPER SIRNA RNAI MAGE-1 

分 类 号：R73-354[医药卫生—肿瘤]