Construction of recombinant adenovirus vector containing human survivin gene and its expression in dendritic cells / 中国实验血液学杂志

ABSTRACT

The study was aimed to construct the recombinant adenovirus vectors containing human survivin gene, and to investigate their expression in transfected dendritic cells. Full length cDNA encoding survivin was obtained by PCR amplification from plasmid pcDNA3.0-survivin. The PCR product was restricted, and then inserted into pShuttle-CMV. The plasmids of pShuttle-CMV-survivin were linearized with PmeI, and the fragment containing survivin was ligated with pShuttle-CMV and transfected into E. coli BJ5183. After homologous recombination in bacteria, the extracted plasmid from the positive bacteria were linearized with PacI, transfected into HEK293 cells with liposome Lipofectamine 2000. Then, the harvested adenovirus supernatants were transfected into dendritic cells. The results showed that the recombinant adenovirus-survivin was constructed successfully and its titer was about 2.65 x 10(9) pfu/ml. The expression of survivin in transfected dendritic cells was confirmed by Western blot analysis. It is concluded that the recombinant adenovirus vector containing human survivin was constructed successfully, which may provide preliminary laboratory evidence for anti-leukemia immunotherapy.