Construction of the recombinant adenovirus vector carrying antisense multidrug resistance-associated protein with AdEasy system

Objective: AdEasy system was used for generating recombinant adenovirus vector carrying antisense MRP in the gene therapy of hepatocellular carcinoma to overcome drug resistance. Methods: The fragment of MRP gene encoding 5'region was cloned reversely into the shuttle plasmid pAdTrack-CMV, with the resultant plasmid and the backbone plasmid pAdEasy-1,the homologous recombination took place in the E.coli BJ5183 and the recombinant adenoviral plasmid was generated. The adenoviruses were packaged and amplified in the 293 cells. Then the viral titer was checked by GFP. Results: The recombinant adenovirus vector carrying antisense MRP was constructed successfully. The viral titer was 2.5 × 109efu/ml,and more than 90% SMMC-7721/ADM cells could be transfected when the MOI was 100. Conclusion: Recombinant adenoviruses with high titer and efficient transfection of HCC cells could be obtained quickly and simply by using AdEasy system, which would provide experimental basis for the mechanisms and reversal methods of the multidrug resistance in human hepatocellular carcinoma.