Dissertation > Excellent graduate degree dissertation topics show

Study on the Generation of Transgenic Embryos of Cattle and Mouse with Lentiviral Vector

Author: SunKeNing
Tutor: LinFeng
School: Henan Agricultural University
Course: Animal Genetic Breeding and Reproduction
Keywords: Lentiviral Vectors Sperm Mediated Lentivirus Packaging Transgene
CLC: S814.8
Type: Master's thesis
Year: 2011
Downloads: 4
Quote: 0
Read: Download Dissertation

Abstract


Lentiviral vectors were derived from HIV-1 provirus genome. Lentiviral vectors are a type of retrovirus that can infect both dividing and nondividing cells. Lentiviruses can make long-term stability of foreign genes expression. Lentiviral vectors have been successfully applied to a variety of transgenic animal production. This study uses a three-plasmid lentiviral vector system, it is self-inactivated vector and containing WPRE and other regulatory elements.Fat-1 gene is derived from the Caenorhabditis elegans, and its expression product is a type ofω-3 fatty acid desaturase, It can changeω-6 polyunsaturated fatty acids (polyunsaturated fatty acids, PUFAs) intoω-3 PUFAs, improve the level ofω-3 PUFAs.ω-3 PUFAs plays an important role in keeping healthy for human and has significant functions for prevention and treatment for many human illnesses.This study constructed a lentiviral vector that can express Fat-1 and the marker gene EGFP mediated by IRES. It laid the foundation for the subsequent production of transgenic animals.The efficiency of current lentivirus packaging is low and unstable. To improve the efficiency of lentivirus packaging, the influences of state of cell growth and existence on virus packaging were studied; the effects of adherent and suspended 293T cells using calcium phosphate method on packaging lentivirus were compared. The results show that the titer of lentivirus packaged with suspension cells was nearly 3.5 folds higher than that packaged with adherent cells. Moreover, two advantages, higher repeatability and less cell culture time which was at least 24 h shorter, of lentivirus packaging using suspension cells directly were also showed in this study. Therefore, a highly efficient technology of lentivirus packaging using suspended cells was established in our study.Lentivirus were incubated with sperm, then transgenic animals were produced with in vitro fertilization. We did not obtain any positive embryos in mice and cattle with this method. In order to study the influence of fertilization on the infection of embryos by lentivirus, lentivirus of low titer (106 TU / ml) were microinjected into perivitelline spaces of mouse fertilized eggs and oocytes. The positive rate of the group of fertilized eggs was 8.3%, group of oocytes did not have positive embryos. The results show that the process of fertilization did not increase the efficiency of lentivirus infection on mice embryos.In order to study lentiviral vectors in the application of transgenic cows, perivitelline spaces of oocytes, zygotes, 2- to 4-cell embryos, 8- to 16-cell embryos and morulas were microinjected with of 108 TU / ml. Results: groups of 2- to 4-cell embryos and zygotes had not any positive embryos; the positive rate of the group of oocytes was 33.3%; the positive rate of the group of 8- to 16-cell embryos was 2.2%; the positive rate of the group of morula was 71.4%. The results showed that microinjecte lentivirus into perivitelline spaces of oocytes is a very efficient way to produce transgenic cattles. When bovine embryos reach the 8- to 16-cell stage, with the increase of genome transcription activity, the efficiency of lentiviral infection is increasing.

Related Dissertations

  1. Resistance Analysis of the Code Region of Pib Gene to Blast in Transgenic Rice under Different Promoters,S435.111.41
  2. Cloning and Characterization of Photoperiod Sensitive Gene ZmELF4 in Maize,S513
  3. Study on Gene Therapy of SiRNA Targeted at Telomerase of Hepatocellular Carcinoma Transduced by Lentivirus in Vitro,R735.7
  4. Construction of Multiple Gene Coexpression Lentiviral Vector System Using GFP as the Reporter Gene,R346
  5. Establishment of the Dual-Luciferase Reporter System for Screening Anti-Tumor Medicine Base on the P21 Pathway,R96
  6. The Construction of CD8α + Antigen Cloning Vector and Expression in Dendritic Cells,R587.1
  7. The Transformation of Genes Related to Glycine Betaine Biosynthesis from Salicornia Europaea Improves Salt-tolerance of Populus×Eurnmericana and Tobacco,S572
  8. Establishment of CYP3A11 Gene Knock-down Mouse by Lentiviral Transgenesis,Q78
  9. Bidirectional Adjustment of Manganese Superoxide Dismutase Overexpression on Esophageal Carcinoma TE-1 Cell,R735.1
  10. RNAi inhibition of Smad6 and under the action of TGF-β1 in mouse bone marrow-derived dendritic cells changes of biological characteristics,R392
  11. Exprimental Study on Differentiation of Rat Bone Marrow Mesenchymal Stem Cells Modified by Basic Fibroblast Growth Factor in Vitro,R329
  12. Study on Inhibiting Bombyx Mori Baculovirus from Multiplication Basded on Transgenic RNAi,Q78
  13. Preliminary Study on Transgenic Misgurnus Anguillicaudatus with PiggyBac,Q78
  14. Stability of Transgene in Micropropogation Plants of Transgenic Birch,S792.15
  15. Cloning and Expressing Regulation of Sucrose Non-fermenting-1-related Protein Kinase-1 (SnRK1) from Pingyitiancha (Malus Hupehensis Rehd.) and Researching of Its Function,S661.1
  16. Construction of an Expression Vector and Screening a Stable Transfection Cell Line for HSF4b Protein,R776.1
  17. Studies of Construction of Overexpression Vectors Containing RubisCO Activase Gene (RCA) and Genetic Transformation,S642.2
  18. Turn human growth hormone (hGH) gene F_4 generation the red carp transit plant gene mapping studies,Q789
  19. The Study on Mechanism and Interfere Factors of Parkinson’s Disease,R742.5
  20. Expression of cptpi Gene from Spinach and cfba Gene from Rice in Anabaena sp. PCC 7120 and Its Regulation of Photosynthesis,Q945.11

CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > General Animal Science > Livestock and poultry breeding > Reproductive biotechnology
© 2012 www.DissertationTopic.Net  Mobile