[Construction of recombinant adenovirus vector containing human survivin gene and its expression in dendritic cells].
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.
Zhu
Xiong-Peng
XP
Fujian Institute of Hematology, Union Hospital, Fujian Medical University, Fuzhou 350001, China. xiongpengzhu@163.com
Chen
Zhi-Zhe
ZZ
Lin
Xu
X
Hu
Jian-Da
JD
Li
Chun-Tuan
CT
Yang
Ting
T
Xu
Zheng-Shu
ZS
Lü
Lu-Lu
LL
Chen
Cai-Ping
CP
Zhang
Lang-Hui
LH
D000256:Adenoviridae / Q000235:genetics / Q000378:metabolism*; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D005822:Genetic Vectors; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins / Q000096:biosynthesis* / Q000235:genetics; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D000077022:Survivin; D014162:Transfection
C107489:BIRC5 protein, human; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D011994:Recombinant Proteins; D000077022:Survivin