Dissertation > Excellent graduate degree dissertation topics show

The Construction of a Novel Vector Targeting to the hrDNA Locus with VEGF165 and Targeted Expression of VEGF165 in Cell

Author: LiangZongMin
Tutor: HuDongZuo
School: Central South University
Course: Surgery
Keywords: Human gene vector VEGF165 Site-specific integration
CLC: R346
Type: PhD thesis
Year: 2007
Downloads: 61
Quote: 0
Read: Download Dissertation

Abstract


Vascular endothelial growth factor (VEGF) is an angiogenicregulator to stimulate endothelial cell migration, proliferation andangiogenesis. VEGF gene therapy is a new promising approach to inducetherapeutic angiogenesis for the treatment of ischemic myocardial andlimb diseases. Recently, Clinical studies have demonstrated successfuloutcomes using plasmids, retroviruses, and adenoviruses.Selecting vector is one of the important factors in gene therapy.Indeed, among the universe of clinical gene transfer trials, the viral vectoris widely used, including retrovirus, adenovirus, or adenovirus associatedvirus. However, the safety of those vectors is poor and made it a seriousconcern. Besides immunogenicity caused by viral vectors, a furtherproblem with viral approaches is that viral vectors show a preference tointegrate into the transcription or control region of active genes, then itmay induce tumour and other disorders. Efficient and safe nonviralvectors have become of the subject many researchers pursued. Thus, theresearchers of National lab of medical genetics of China have constructeda new nonviral vector—hrDNA targeting vector: pHrneo, pHrneo is ahuman source gene vector that targets heterologous gene to the humanribosomal DNA (hrDNA)locus with an efficiency of integration up to10-4-10-5. In this study, pHrneo was chosen as the vector in the research of gene therapy with VEGF165. Human ribosomal RNA gene (rDNA)locates in D and G group chromosome. In human cells, there are about600-800 copies rDNA per cell. For the specific structure of rDNA, anexogenous gene inserting into ribosomal gene target site to destroy a copyof rDNA, there may be no effect on the cell physiological function.After amplified by PCR, the VEGF165 fragment, together withCMV, EGFP and BGHploy(A), was combined to construct a expressioncassette. Then the cassette was inserted into pHrneo vector to constructpHrneo-VEGF165. Through electroporation pHrneo-VEGF165 wastransferred into HT1080 cells. Finally we identified the homologousrecombinants by PCR, and tested the expression of VEGF165 in thehrDNA locus.The VEGF165 was successfully targeted into hrDNA locus ofHT-1080 by pHrneo-VEGF165, and the efficiency of site-specificintegration was 7.7×10-5 The quantity of VEGF165 fusion protein was(1000±23) pg·106 cells-1·24 h-1 tested by ELISA, the mRNA and fusionprotein of VEGF165 were detected by RT-PCR and Western-blottingrespectively. This research will provide new insights into the clinicalresearch of ischemic limb and myocardial diseases.

Related Dissertations

  1. The Research of FAK and Akt Signal Pathways in HUVEC after Vegf121 or VEGF165 Isoforms Treatment in Vitro,R758.2
  2. The Study of Superparamagnetic Chitosan (pDsVEGF165)gelatin Microsoheres for the Cranial Defects,R318.08
  3. The Effects of 17β-estradiol in Rat Liver Fibrosis,R575.2
  4. The study and application of the Rep protein -mediated site-specific integration system of cis-elements,Q75
  5. Research of EPC Change with Coronary Heart Diseases Patients and Effect of LPS and Rosiglitazion on EPC,R541.4
  6. Revascularization of Islet Grafts Induced by Rat Vascular Endothelial Cell Transfected by VEGF165,R587.1
  7. Targeted Genetic Transformation in Cultured Silkworm Cells Based on PiggyBac Transposon,Q78
  8. Tissue-type Plasminogen Activator Up-regulate Expression of Vascular Endothelial Growth Factor in ECV304 Cells Via ERK and P38 Signaling Pathway,R654.4
  9. Construction of a Mouse Model Expressing φC31 Integrase,Q78
  10. VEGF165 and Angiopoietin-1 in ischemic myocardium -specific expression treatment of acute myocardial infarction study,R542.22
  11. AAV-2 P5 promoter function of the molecular basis of,Q75
  12. Preliminary Study of Correction of Murine Hemophilia B by Hydrodynamic Gene Dlivery and Site-specific Genomic Integration,R554.1
  13. Site-specific Integration of Heterologous Gene into Bacillus Thuringiensis Chromosome and Its Expression,Q78
  14. MLC-2v cardiac-specific core promoter and RTP801 sexual enhancer core hypoxia response under the control of VEGF_ (165) and FGF_2 efficient coexpression,R363
  15. Effect of VEGF165 on the Permeability of Blood Brain Barrier after Intracerebral Hemorrhage,R743.3
  16. p53 Anti-tumor Research in Bel-7402 by Using Human-derived Vector,Q789
  17. The Construction of Histone Acetyltransfarese Modified Human-Derived Vectors and Their Function Study,Q78
  18. Study of a Novel Nanosphere Complexes-mediated VEGF Gene Transfection on Fibroblasts in Vitro,R346
  19. Construction of AAV Co-expression System for Human Angiopoietin1 and VEGF Gene,R346
  20. Construction of Microencapsulated Cell Expressing Human Vascular Endothelial Growth Factor,R329

CLC: > Medicine, health > Basic Medical > Human biochemistry, molecular biology
© 2012 www.DissertationTopic.Net  Mobile