2. TERMINOLOGY
Transfection : Introduction of foreign DNA into
eukaryotic cells usually animal cells.
Transfectants: Cells that have incorporated foreign
DNA.
Stable : Integrated foreign DNA.
Transient: Does not integrate foreign DNA, but genes
are expressed briefly.
3.
4. DIFFERENCE
Cloning (uptake of genetic material) Cancer
Transformation Transfection Transformation
In unicellular
organisms like
prokaryotes (bacteria)
or unicellular
eukaryotes (amoeba)
In Metazoan
Eukaryotic Cells
Advancement of a
metazoan eukaryotic
cell from being non-
cancerous to
cancerous
6. VIRUSES IN USE
Viral Vector DNA Insert
Size
Expression Pitfalls
Retro viral 8 kb Stable Random insertion site
Lenti viral 9 kb Stable Random insertion site
Adeno Virus 8 kb Transient Highly immunogenic
Adeno associated
Virus
5 kb Stable, site
specific location
Requires helper virus
and difficult to remove
Herpes Simplex
Virus
30-40 kb Transient No gene expression
during latent infection
Vaccina Virus 25 kb Transient Potential cytopathic
effects
7. FOOT NOTES
100% efficiency
Toxicity
Strong immune response
Untargeted integration of genes
Complexity of generating recombinant viruses
Limited packaging capabilities
In vivo DNA delivery
8. VIRUS LIKE PARTICLES(VLP)
Alternative approach to classical methods
Viral capsid- without viral genetic information.
Eg: Pappilloma viruses: L1 and L2 proteins
Predominantly use – vaccination
Gene delivery – human polymo JC virus, murine
polymovirus, pappilomaviruses and AAV- based VLPs.
Isolation and
purification of
viral capsid
proteins
Empty viral
particles
reconstituted and
stored at -80 0C
Packaging with
DNA or siRNA
inside empty viral
particles
9. SINGLE WALLED CARBON NANOTUBES –
MECHANICAL METHOD
Unidimensional layer of carbon-hexagons form a
tube.
Functionalized- amino or carboxyl group
Covalent or non-covalent bond with biomolecules.
Diameter: 1-5nm; Length: 50-200nm
Success: In vivo siRNA delivery.
13. SUMMARY
Chemical methods : system needs to be adapted to
the cargo to be delivered.
No separate genetic protocols for siRNA and
plasmid DNA delivery.
Physical methods : cytotoxicity, cellular uptake
insufficient mostly.
What is needed?
Specific tailor-made DNA and siRNA delivery systems
Nucleic acid-based therapeutics : individualized medicines for
specific disease variation.