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- W2361686799 abstract "Objective To construct a recombinant adenovirus for expression of human TNF-related apoptosis inducing ligand(hTRAIL) gene.Methods Amplify gene fragment by PCR using the TRAIL gene extracted from recombinant plasmid pThioHisA-TRAIL as a template,insert into shuttle plasmid pShuttle-CMV and transform to E.coli DH 5α.Screen recombinant plasmid pShuttle-CMV-TRAIL and transform to competent BJ5183 cells previously transformed with pAdEasy-1.Screen recombinant plasmid pAdEasy-TRAIL and transfect to 293 cells in mediation of Lipofectamine to prepare replication-deficient adenovirus Ad-TRAIL carrying full-length of hTRAIL gene.Purify the prepared Ad-TRAIL virus particles by density gradient centrifugation with cesium chloride and determine the titer.Determine the transcription of TRAIL gene in 293 cells by RT-PCR.Transfect YTMLC cells with Ad-TRAIL and determine the expression of TRAIL by IFA and flow cytometry.Results The virus particle count and titer of purified Ad-TRAIL were 2.77×1012 VP/ml and 109.5(3.2×109)CCID50/ml respectively.The gene fragment at a length of about 843 bp was amplified by RT-PCR from transfected 293 cells,which was consistent with that expected.Both flow cytometry and IFA proved the expression of TRAIL protein in YTMLC cells,and the expressed product mainly existed in cytoplasm.Conclusion Recombinant adenovirus Ad-TRAIL for expression of hTRAIL was successfully constructed,which provided a basis for gene therapy of tumors." @default.
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- W2361686799 date "2007-01-01" @default.
- W2361686799 modified "2023-09-26" @default.
- W2361686799 title "Construction and Identification of Recombinant Adenovirus for Expression of hTRAIL" @default.
- W2361686799 hasPublicationYear "2007" @default.
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