Transcript Molecular Cloning Methods

RECOMBINANT DNA TECHNOLOGY
PART 2
Topics
• Genomic libraries
• Id a specific clone or sequence within a library
• Transgenic plants
• Transgenic animals
• Timely topics
Genomic library contains all of the organism’s genome
Probe design
Bacterial clones and finding desired clone
Probe des
A Final
about vectors : There are many types
• Small vectors
• Large vectors
• Expression vectors
• Viral vectors

Transgenic Mice
transgen
Non-homologous recombination
Linear DNA
Knockout Mice
G418
gancyclovir
Homozygous
knockout
Phenotype of the knockout mouse gives information on protein function
Applications of DNA technology
• Agriculture
• Drug design
• Genetic testing
• Gene therapy
Genetic Modified Plants: “GM”
Tobacco with luciferase gene from firefly
Transgenic plant
Tumor Inducing Plasmid
Model for making genetically modified (GM) plants
GM crops
• Bt toxin genes (Bacillus thuringensis)
Bt Toxin
Cry toxins and cry genes: damages gut of larvae
GM Plants
• Roundup  ready plants: herbicide resistant
• Herbicide known as: Glyphosate
Glyphosate
• Enzyme inhibitor
• Prevents amino acid synthesis
• Prevents weed growth
• Environmental concerns
Antisense Technology
•
5´ C A U G 3´
3´ G U A C 5´
mRNA
Antisense RNA
Anti-Sense Technology
&
FLAVR SAVRTM Tomato
•Anti-sense DNA technology inhibits polygalacturonase (PG)
The Flavr-Savr Tomato
• Ripens on the vine
• Preserves firmness and flavor
• Uses anti-sense technology
Cloning of a whole organism: Reproductive Cloning
SHEEP CELLS
54
27
Somatic Cell
Germ Cell
For ex. Mammary cell
ovum
How to process begins:
54
27
54
discarded
Somatic Cell
Germ Cell
For ex. Mammary cell
ovum
Now the germ cell contains 54 chromosomes!
• The germ is stimulated “electrically”.
• Cell division in some instances begins.
• Individual is identical to host “DNA”
donor.
• Aging affects?
“Was Dolly a sheep in lamb’s clothing?”
Somatic Gene Therapy
Successes
• SCIDS (Severe Combined Immunodeficiency Disease)
• Head and neck cancers
But also failures and controversy