摘要
目的构建基质金属蛋白酶-26(MMP-26)小干扰RNA(siRNA)表达质粒,以研究其对人肿瘤细胞MMP-26基因表达的沉默效果,探索肿瘤基因治疗的新途径。方法根据MMP-26基因mRNA序列,设计有小发夹结构的3条寡核苷酸序列,克隆到空载体pSUPERIOR.puro中,构建重组质粒,同时设计构建分别针对GFP和-βactin的干扰质粒作为阴性对照和阳性对照,并进行酶切鉴定和测序分析。结果酶切及测序证实重组质粒构建成功。结论利用RNA干扰(RNAi)技术可成功构建siRNA表达载体。
Objective To establish the recombinant plasmied of small interfering RNA （siRNA） against matrix metalloproteinase- 26 （ MMP-26）, observe dumbness effect of the MMP-26 expression in human carcinoma, and explore the new method of gene therapy. Methods Recombinant were designed and established by targeting gene MMP-26 and plamid pSUPERIOR, puro based on MMP-26 mRNA sequence. Three pairs of oligonucleotides were synthesized and inserted into plamid pSUPERIOR, puro to generate siRNA eukaryotic expression vectors. DH5α strains were transformaed,, plasmid were extracted, and the recombinant sequences were identified. Interfering plasmid from GFP and β-actin were established as negative and positive controls for MMP-26. Results The result of recombinant sequence was the same as aim sequence. The recombinant vectors were established successfully. Conclusion siRNA reeombinant can be established successfully by RNA interference （RNAi） technique.
出处
《中国实验诊断学》
2006年第10期1135-1137,共3页
Chinese Journal of Laboratory Diagnosis
基金
国家自然科学基金资助项目(30371656)
