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Transgenesis

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Title: Transgenesis  
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Subject: Genetic engineering, Amflora, Plant tissue culture, Viruses/Did you know/7, Genome engineering
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Transgenesis

Transgenesis is the process of introducing an offspring. Transgenesis can be facilitated by liposomes, plasmid vectors, viral vectors, pronuclear injection, protoplast fusion, and ballistic DNA injection.

"super mice" of the 1980s. These mice were able to produce the human protein tPA to treat blood clots.

Contents

  • Using plasmids from bacteria 1
  • Gene transfer technology 2
    • DNA microinjection 2.1
    • Retrovirus-mediated gene transfer 2.2
    • Stem cell transgenesis 2.3
      • Multipotent stem cell transgenesis 2.3.1
      • Pluripotent stem cell transgenesis 2.3.2
      • Totipotent stem cell transgenesis 2.3.3
  • Applications 3
    • Pharming 3.1
    • Medical 3.2
  • Ethical Concerns 4
  • Diagram 5
  • References 6

Using plasmids from bacteria

The most common type of transgenesis research is done with bacteria and viruses which are able to replicate foreign DNA.[1] The plasmid DNA is cut using restriction enzymes, while the DNA to be copied is also cut with the same restriction enzyme, producing complementary sticky-ends. This allows the foreign DNA to hybridise with the plasmid DNA and be sealed by DNA ligase enzyme, creating a genetic code not normally found in nature. Altered DNA is inserted into plasmids for replication.[2]

Gene transfer technology

DNA microinjection

The Desired gene construct is injected in the pronucleus of a reproductive cell using a glass needle around 0.5 to 5 micrometers in diameter. The manipulated cell is cultured in vitro to develop to a specific embryonic phase, is then transferred to a recipient female. DNA microinjection does not have a high success rate (roughly 2% of all injected subjects), even if the new DNA is incorporated in the genome, if it is not accepted by the germ-line the new traits will not appear in their offspring. If DNA is injected in multiple sites the chances of over-expression increase.[3]

Retrovirus-mediated gene transfer

A homozygous genetic offspring are born.[3]

Stem cell transgenesis

Multipotent stem cell transgenesis

Multipotent stem cells can only differentiate into a limited number of therapeutically useful cell types, nevertheless their safety and relative lack of complexity to us have resulted in the vast majority of current personalized cellular therapeutics involving multipotent stem cells (typically mesenchymal stem cells from adipose tissue).[4]

Pluripotent stem cell transgenesis

Transgenic vectors can be delivered randomly, or targeted to a specific genomic location, such as a safe harbor . Scientists have performed research and technology development to provide the tools necessary to permit safe and effective pluripotent stem cell (PSC) transgenesis.[5][6][7][8][9][10][11][12]

Totipotent stem cell transgenesis

The manipulated gene construct is inserted into totipotent stem cells, cells which can develop into any specialized cell. Cells containing the desired DNA are incorporated into the host’s embryo, resulting in a chimeric animal. Unlike the other two methods of injection which require live transgenic offspring for testing, embryonic cell transfer can be tested at the cell stage.

Applications

Pharming

Pharming is a biotechnology since the development of trangsgenic "super mice" in 1982. "Super mice" were genetically altered to produce the human drug, tPA (tissue plasminogen activator to treat blood clots), in 1987.[2] Since then "super mice" pharming has come a long way. Using RNA interference scientists have produced a cow whose milk contains increased amounts of casein, a protein used to make cheese and other foods, and almost no beta-lactoglobulin, a component in milk whey protein that causes allergies.[13]

"Pharming Examples:"[14]

  • Haemoglobin as a blood substitute
  • human protein C anticoagulant
  • alpha-1 antitrypsin (AAT) for treatment of AAT deficiency
  • insulin for diabetes treatment
  • vaccines (antigens)
  • growth hormones for treatment of deficiencies
  • factor VIII blood clotting factor
  • factor IX blood clotting factor
  • fibrinogen blood clotting factor
  • lactoferrin as an infant formula additive

Medical

Transgenesis can be used to neutralize genes that would normally prevent

  1. ^ "Mousepox Case Study — Module 4.0".  
  2. ^ a b Redway, Keith. "Transgenic organisms". Gene Manipulation & Recombinant DNA.  
  3. ^ a b Margawati, Endang Tri (January 2003). "Transgenic Animals: Their Benefits To Human Welfare".  
  4. ^ clinicaltrials.org
  5. ^ Capecchi MR (June 2005). "Gene targeting in mice: functional analysis of the mammalian genome for the twenty-first century". Nat. Rev. Genet. 6 (6): 507–12.  
  6. ^ Cong L, Ran FA, Cox D, et al. (February 2013). "Multiplex genome engineering using CRISPR/Cas systems". Science 339 (6121): 819–23.  
  7. ^ DiCarlo JE, Norville JE, Mali P, Rios X, Aach J, Church GM (April 2013). using CRISPR-Cas systems"Saccharomyces cerevisiae"Genome engineering in . Nucleic Acids Res. 41 (7): 4336–43.  
  8. ^ Friedland AE, Tzur YB, Esvelt KM, Colaiácovo MP, Church GM, Calarco JA (August 2013). via a CRISPR-Cas9 system"C. elegans"Heritable genome editing in . Nat. Methods 10 (8): 741–3.  
  9. ^ Hwang WY, Fu Y, Reyon D, et al. (March 2013). "Efficient genome editing in zebrafish using a CRISPR-Cas system". Nat. Biotechnol. 31 (3): 227–9.  
  10. ^ Nguyen HN, Reijo Pera RA (2008). "Metaphase spreads and spectral karyotyping of human embryonic stem cells". CSH Protoc: pdb.prot5047.  
  11. ^ Mali P, Yang L, Esvelt KM, et al. (February 2013). "RNA-guided human genome engineering via Cas9". Science 339 (6121): 823–6.  
  12. ^ Xue H, Wu J, Li S, Rao MS, Liu Y (March 2014). "Genetic Modification in Human Pluripotent Stem Cells by Homologous Recombination and CRISPR/Cas9 System". Methods Mol. Biol.  
  13. ^ Lopatto, Elizabeth (October 1, 2012). "Gene-Modified Cow Makes Milk Rich in Protein, Study Finds".  
  14. ^ Buy M (1997). "Transgenic Animals". CCAC Resource Supplement. Canadian Council on Animal Care (CCAC). 
  15. ^ "Actionbioscience | Transgenic Animals: Their Benefits To Human Welfare". actionbioscience.org. Retrieved November 29, 2014. 
  16. ^ "Actionbioscience | Ethical Issues in Genetic Engineering and Transgenics". actionbioscience.org. Retrieved November 29, 2014. 

References

Note: New genotypes created with transgenic technologies also require multiple backcrossings. Furthermore, backcrossing does not account for the majority of time required to create, field test and release/commercialize a new variety.

.
A diagram comparing the genetic changes achieved through conventional plant breeding, transgenesis and cisgenesis

Diagram

Transgenesis has created certain ethical concerns. Examples include rights for animals that have been improved intellectually, legal ramifications, and possible health risks.[16]

Ethical Concerns

[15]

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