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Construction of Tropism Liver Cells Recombinant Adenoviral Vector for Gene Therapy

Author: TangDan
Tutor: ZhangMingMan
School: Chongqing Medical University
Course: Pediatrics
Keywords: Adenovirus Hepatitis B virus Targeting Gene therapy
CLC: R346
Type: Master's thesis
Year: 2010
Downloads: 27
Quote: 0
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Abstract


The Objective: recombinant adenovirus ( RAD ) is commonly used gene therapy vector , but it lacks Hepatotropism , the subject to be constructed through the transformation of the rAd cilia structure , has Hepatotropism RAD carrier , targeted gene therapy for liver disease . AdEasy System : In the currently used rAd high expression vector system , based on the capsid protein of hepatitis B virus (HBV) having the natural peptides Hepatotropism PreS1 the gene encoding 21-47 amino acid at position ( PreS1 ) the designated clones to in AdEasy System skeleton the plasmid pAdEasy - 1 of the HI Loop coding gene loci , constructed recombinant skeleton the plasmid pAdEasy - 1 - anti-preS1 then pAdEasy -1 - PreS1 with shuttle plasmid pShuttle - IRES - hrGFP - 1 homologous recombination composed the viral plasmid Pad - preSl packaging to generate adenoviral gene therapy vectors RAD - preS1 after transfected HEK293 cells . 1 results: The cloned the preS1 gene fragment adenovirus recombinants by gene sequencing revealed the correct sequence ; viral plasmid transfected HEK293 cells , a week after the fluorescence observation found that the expression of a fluorescent protein , suggesting that adenovirus generation ; recombinant adenovirus gene can to extract Hepatotropism gene fragment preS1 template for PCR detection ; extraction recombinant adenovirus protein by immunoblotting can detected the expression of the target protein PreS1 ( 21-47 ) , suggesting that adenovirus gene therapy vector rad the - PreS1 generation ; the build of RAD - PreS1 of infection a variety of cells , the virus titer relative addicted to the liver cells sex . Conclusion: the the rAd cilia ball domain HI Loop coding loci addicted to the liver fragment PreS1 of 21-47 amino acids into the HBV surface protein encoding gene preS1 packaging HEK293 cells to generate the recombinant adenovirus gene therapy vector rAd - PreS1 , by in vitro experiments show that the liver cells having a relatively isotropic addicted .

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CLC: > Medicine, health > Basic Medical > Human biochemistry, molecular biology
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