特异切割多药耐药基因(MDR1)的核酶设计与克隆  

Designing and Cloning of a Hammerhead Ribozyme which can Site-specifically Cleave Human MDR1?mRNA

作　　者：刘顺英[1] 孙宁[1] 武景阳[2] 单祥年[2] 

机构地区：[1]南京铁道医学院附属医院消化科,南京210009 [2]南京铁道医学院附属医院生物学教研室,南京210009

出　　处：《南京铁道医学院学报》1999年第4期225-228,共4页Journal of Nanjing Railway Medical College

基　　金：铁道部科技基金!资助课题(96 - J- 2)

摘　　要：目的:设计与克隆特异切割多药耐药基因(MDR1) 的核酶。方法:针对人类MDR1 mRNA第Ⅶ外显子附近第196 密码子的GUC 序列,按照“锤头结构”模型,设计、合成核酶,并定向克隆入载体PGEM-3Zf(t) 中;通过PCR- SSCP法及限制性分析,确保扩增片断为外源插入的DNA序列,并经DNA测序证实。结果:重组质粒插入片断序列与设计序列一致。结论:设计、克隆核酶作用人类MDR1Objective:This study was aimed to generate a ribozyme capable of specific cleavage of MDR1 mRNA.Methods:Based on “hammerhead model”,the ribozyme was designed to cleave the GUC sequence in coden 196 of MDR1 mRNA.The synthesized ribozyme sequence was cloned into pGEM-3Zf(t).The resulting clones were screened for presence of DNA insert by PCR-SSCP and restriction analysis and further confirmed by DNA sequencing.Results:Recombinent plasmids containing the expected ribozyme sequence with correct orientation were obtained.Conclusion:The procedure described in this paper proves effective in design and cloning of the ribozyme targeted against the RNA transcripts of human MDR1 gene.

关 键 词：多药耐药基因 核酶 分子克隆 肿瘤 

分 类 号：R730.53[医药卫生—肿瘤] Q55[医药卫生—临床医学]