[Construction and expression of recombinant adenovirus expression vector of beta-defensin-2]

Zhuo Shi, Qiang Shu, Zheng-yan Zhao
Zhejiang da Xue Xue Bao. Yi Xue Ban, Journal of Zhejiang University. Medical Sciences 2006, 35 (6): 590-5

OBJECTIVE: To explore the possibility of stable expression of cationic peptide beta-defensin-2 in eukaryocytes with adenovirus vector.

METHODS: The rat beta-defensin-2 (rBD2) gene was cloned at the downstream of CMV promoter of the adenoviral shuttle plasmid pShuttle-CMV. Then pShuttle-CMV-rBD2 was transformed into E. coli BJ5183-AD-1, in which recombination occurred between plasmids and pAdEasy-1 to construct pAdEasy-rBD2. After confirmation by endonuclease, linear pAdEasy-rBD2 was transformed into 292 cells to obtain packaged adenoviral expression vector, which was used to infect cos-7 cells and to establish respiratory adenovirus infection model of rat. The in vivo and in vitro expression activity of recombinant adenovirus was detected by Western blot and immunohistochemistry.

RESULT: The inserted DNA of pShuttle-CMV-rBD2 consisted of rat beta-defensin-2 gene. The pathological effect of infected cells, electronic microscopic observation and PCR showed that the recombinant adenovirus vector was constructed successfully. The concentration of the adenovirus was 10(9) PFU/ml. The vector expressed rat beta-defensin-2 efficiently in vivo and in vitro.

CONCLUSION: The recombinant adenovirus vector can express cationic peptide beta-defensin-2 in eukaryocytes.

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