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 design

A Final about vectors : There are many types

• Small vectors

• Large vectors

• Expression vectors

• Viral vectors

Transgenic Mice

transgene

Linear DNA

Non-homologous recombination

http://spine.rutgers.edu/cellbio/assets/flash/transgen.htm

Knockout Mice

Phenotype of the knockout mouse gives information on protein function

Homozygous knockout

G418gancyclovir

http://www.bio.davidson.edu/courses/genomics/method/homolrecomb.htm

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´     mRNA

3´   G U A C   5´     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

Somatic Cell Germ Cell

For ex. Mammary cell ovum

SHEEP CELLS

54 27

Somatic Cell Germ Cell

For ex. Mammary cell ovum

How to process begins:

542754

discarded

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