10—23脱氧核酶抑制铜绿假单胞菌耐药基因VIM2作用的初步研究

Research of 10-23 deoxyribozymes inhibit VIM2 gene in Pseudomonas aeruginosa

目的探讨10-23脱氧核酶（DRz）抑制铜绿假单胞菌耐药基因VIM2的表达及其在逆转铜绿假单胞菌耐药中的作用。方法根据计算机模拟的VIM2 mRNA的二级结构设计合成5条VIM2特异的10-23DRz（DRzl～DRz5），在无细胞反应体系中鉴定10—23DRz对VIM2 mRNA的切割活性后，将其导入铜绿假单胞菌，利用E—test法检测亚胺培南对处理前后铜绿假单胞菌的MIC值。结果DRz3、DRz4和DRz5在无细胞反应体系中可对VIM2 mRNA进行有效切割，且其活性具有高度特异性。与对照相比，10-23DRz可以降低业胺培南对铜绿假译胞菌的MIC值。结论实验中所设计的10-23DRz能在细胞外高效特异性的切割VIM2 mRNA；在细胞内能协同亚胺培南抑制铜绿假单胞菌耐药。

Objective To investigate the inhibitory effects of 10-23 deoxyribozyme （DRz） on expression of VIM2 and the effect against tolerance of Pseudomonas aeruginosa. Methods Five 10-23DRzs targeting VIM2 gene （ DRz1-DRz5 ） were designed according to the predicted secondary structure of VIM2 mRNA. Their cleavage activity and specificity were identified in cell free conditions. 10-23DRz were introduced into Pseudomonas aeruginosa producing β-lactamas by electroporation procedure. Following electroporation, MICs of imipenem against Pseudomonas aeruginosa were detected by E-test method. Results Three of the five designed 10-23DRz （DRz3-DRz5） could cleavage VIM2 mRNA efficiently and specifically. MIC values aginst Pseudomonas aeruginosa were lower than that in eontrol DNA electroporated. Conclusion 10- 23DRz can cleave VIM2 mRNA with effectiveness and specificity, and exhibit synergic effect with imipenem on Pseudomonas aeruginosa, which will probably play an important role in the gene therapy of tolerance.