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  • Title: [Construction and identification of replication-defective recombinant adinovirus expressing human bone morphogenetic protein 4].
    Author: Wang X, Wang C, Zhao Y.
    Journal: Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi; 2007 Apr; 21(4):396-400. PubMed ID: 17546887.
    Abstract:
    OBJECTIVE: To construct a replication-detective recombinant adinovirus including tne target gene human bone morphogenetic protein 4 (fragment hBMP-4). METHODS: The hBMP-4 gene fragment was cut down from pCS2(+)/hBMP-4, cloned into the eukaryotic expressive vector pcDNA 3.1(+), then subcloned into pShuttle-CMV and transformed into the competent E. coli BJ5183/p by electroporation. The resulting recombinant plasmid pAdE/ hBMP-4 was transformed into the packaging of the cell lines HEK293 to produce the replication-defective recombinant adenoviruses containing the hBMP-4 gene. These replication-defective recombinant adinoviruses were transfected into HEK293 and HeLa cells. Then, total RNA and total protein were detected by RT-PCR and the Western-blot assay. RESULTS: The pAdE/hBMP-4 was confirmed by the restrictional endonuclease digestion. In HEK293 and HeLa cells, the specific transcription of the hBMP-4 gene was confirmed by RT-PCR, and the expression of the hBMP-4 protein was confirmed by the Western-blot assay. CONCLUSION: The replication-defective recombinant adinovirus expression vector containing the hBMP-4 gene can be constructed and expressed successfully, which has laid a foundation for the further research on the genetherapy of hBMP-4.
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