摘要
目的:设计并构建靶向mdr1基因shRNA真核表达质粒。方法:以mdr1为靶基因设计具有短发夹结构的模板寡核苷酸,退火形成互补双链结构,再克隆至pSilencerTM3.1-H1neo载体,构建重组的短发夹shRNA表达载体。采用酶切法和测序法鉴定。结果:经限制性内切酶酶切电泳后显示有66bp模板寡核苷酸片段和4.3kbp的线性化的pSilencerTM3.1-H1neo载体片段。重组子测序结果与Genebank中的mdr1基因cDNA序列相符。结论:重组靶向mdr1基因shRNA表达载体构建正确,为进一步转染耐药的肿瘤细胞逆转肿瘤耐药研究奠定了基础。
Objective：To study the construction of pSilertcerTM 3. 1-H1 neo mdrl short hairpin RNA expression vectors. Methods ：Two different shRNA targets desigened to be homologous to the P-glyeoprotein （P-gp） encoding mdrl mRNA consensus sequence, were annealed and ligated into the BamH Ⅰ and Hind m site of linearized pSilencerTM 3. 1-HI neo vector. The recombinant named as pSilencerTM3. 1-HI neo mdrl-A and mdrl-B shRNA expression plasmids was identificated by restrictive enzyme digestion and sequencing. Results： The fragments of 66 bp and 4. 3 kbp were shown after digestion by BamH I and Hind m and agarose gel electrophoresis. The DNA sequences of pSilencerTM 3. 1-H1 neo mdrl-A and mdrl-B shRNA expression plasmids were proved to be identical to the data of rndrl in Genebank. Conclusions ：pSilencerTM3.1-H1 neo mdrl shRNA expression vectors has been constructed successfully.
出处
《蚌埠医学院学报》
CAS
2007年第4期384-387,共4页
Journal of Bengbu Medical College
基金
国家自然科学基金(No.30171064)
吉林大学创新基金(2003CX045)资助项目
