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- W2369890101 abstract "Objective To construct the replication-deficient recombinant adenovirus carrying vasostatin gene and study its inhibitory effect on pancreatic cancer. Methods A cDNA clone encoding vasostatin was isolated from human skeletal muscle cDNA library by polymerase chain reaction. The shuttle plasmid, pshuttle-CMV-vasostatin, in which the amplified fragment was inserted into the downstream of the cytomegalovirus promoter, was established by ligation. After identification by digestion and sequencing, linearize the shuttle plasmid with Pme I, and then transform the linearized shuttle plasmid into E.coli BJ5183-AdEasy-1 by electroporation. The plasmid was named as pAd-CMV-vasostatin after homologous recombination and identification by kanamycin-selection and restriction digestion. The recombinant plasmid was transfected into cell 293. The recombinant adenovirus was detected by examining the presence of cytopathic effect and identified by polymerase chain reaction. Both RT-PCR and western blotting were used to identify the expression of vasostatin. The xenografted nude mice with pancreatic cancer were established to observe the inhibitory effect of vasostatin on tumor growth. The volume of xenografts was measured every three days during the 3-week treatment. The weight of xenografts was measured at the end of the 21st day. Results The PCR product was about 560 bp confirmed by agarose gel electrophoresis. The successful cloning of vasostatin into pshuttle-CMV was confirmed by restriction endonuclease analysis, and the gene of interest was identified as vasostatin by nucleotides sequencing technique. The recombinants were selected for kanamycin resistance and the recombination was confirmed by the presence of 4.5kb fragment in the restriction endonuclease analysis. The cytopathic effect present after 7 days in cell 293 transfected with linearized pAd-CMV-vasostatin. Both the cytopathic effect of cell 293 and the 560 bp PCR producet indicated the presence of the recombinant adenovirus containing vasostatin gene. The expression of vasostatin was identified by RT-PCR and western blotting. The volume and weight of xenografts in pAd-CMV-vasostatin group were significantly smaller than those in pAd-CMV-LacZ group and PBS group. Conclusion The recombinant adenovirus containing vasostatin gene was constructed successfully, and could significantly inhibit the growth of xenografted pancreatic cancer in nude mice." @default.
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- W2369890101 date "2004-01-01" @default.
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- W2369890101 title "Construction of replication-deficient recombinant adenovirus carrying vasostatin gene and its inhibitory effect on pancreatic cancer" @default.
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