Download Transgenic Organisms

Transgenic Organisms
• What is DNA?
• What do genes do?
• What are restriction enzymes?
• How are transgenic organisms made?
• What can be done with transgenic organisms?
• Gene libraries/Genetic testing
• Genetically-modified crops and livestock
• Protein-producing bacteria
• Gene therapy
What is DNA?
• The cell’s nucleus contains chromosomes (46
chromosomes in human cells).
• Each chromosome is a molecule of DNA.
• Each molecule of DNA is like a cookbook of
protein recipes.
• The recipes are written using a fournucleotide (four letter) alphabet.
• DNA is double-stranded. Each recipe is on
one strand (the other strand is like a back-up).
What is do genes do?
Valine instead of
Glutamate
Sickle-Cell Hemoglobin
• If a chromosome is a cookbook, each gene
is one protein recipe within that cookbook.
Normal Hemoglobin
• Small changes in a recipe can lead to big
changes in the protein that is produced.
• A change of one nucleotide out of around
900 nucleotides changes the hemoglobin
protein from normal to the sickle-cell form!
(That’s like one typo in a 3-page paper).
2 Alpha Chains (141 amino acids each)
2 Beta Chains (146 amino acids each)
2 Alpha Chains (141 amino acids each)
2 Beta Chains (145 normal amino acids, 1 changed amino acid)
What are Restriction Enzymes?
• DNA is double-stranded. The nucleotides in the two strands are complementary: A
pairs with T and G pairs with C.
• Enzymes, made by bacteria, that recognize specific DNA nucleotide sequences, and
make single-stranded cuts in the DNA.
5’
3’
AACTGAATTCCGGATCCGACTAGAATTCATCT
TTGACTTAAGGCCTAGGCTGATCTTAAGTAGA
3’
5’
EcoRI
Recognizes
GAATTC
Bam
HI
Recognizes
GGATCC
Restriction Enzymes: EcoRI as an example
• There are dozens of restriction enzymes now available. Each has a unique
recognition sequence and cut site.
• There are two EcoRI recognition sites in the DNA sequence that is shown.
• The EcoRI recognition sequence is a palindrome (the sequence is
5’ GAATTC 3’ on both strands).
5’
3’
AACTGAATTCCGGATCCGACTAGAATTCATCT
TTGACTTAAGGCCTAGGCTGATCTTAAGTAGA
3’
5’
EcoRI
Recognizes
GAATTC
Bam
HI
Recognizes
GGATCC
Restriction Enzyme: EcoRI Active Site
• The EcoRI active site binds to DNA where ever 5’ GAATTC 3’ occurs.
This works with any DNA.
EcoRI
EcoRI
EcoRI
5’
3’
EcoRI
AACTGAATTCCGGATCCGACTAGAATTCATCT
TTGACTTAAGGCCTAGGCTGATCTTAAGTAGA
3’
5’
EcoRI
Recognizes
GAATTC
Bam
HI
Recognizes
GGATCC
Restriction Enzymes: EcoRI Endonuclease Activity
• EcoRI cuts the sugar-phosphate backbone between the G and A in the
regognition sequence GAATC.
• The cuts in the two strands are off-set from one another. This leaves short
single-stranded ends. The ends are called sticky ends because they will
stick to each other (they have complementary sequence).
EcoRI
Recognizes
GAATTC
Bam
HI
Recognizes
GGATCC
How are transgenic organisms made?
• Plasmids are collected from bacteria. Plasmids are
small bits of DNA that can be taken up and given off
by bacteria. They can be used as vectors for getting
foreign DNA into bacteria.
• Plasmid DNA is collected and cut with a restriction
enzyme.
• DNA of interest (like human DNA) is collected and
cut with the same restriction enzyme. This results in
human DNA and plasmid DNA having
complementary sticky ends.
• Human DNA and plasmid DNA is mixed together
and their sticky ends will bind with one another to
form recombinant plasmids.
• Recombinant plasmids are then mixed with, and
taken up by, bacterial cells. This creates
recombinant (or transgenic or genetically-modified)
organisms.
• The same process can be used to make other types of
transgenic organisms as long as we have an
appropriate vector.
Single-Gene versus Multifactorial Traits
Most complex traits are influenced by
many genes and many environmental
factors. We call these multifactorial
traits. These account for the amazing
variation we see among organisms in the
world.
If all traits were determined by single genes,
this is how the world would look. Thankfully,
that’s not how life is.
We can only modify single genes. Thus, we
only have ability to alter a very limited array of
genetic traits.
What can be done with transgenic organisms:
Gene Libraries/Genetic Tests
• Each of the genes of an organism can be inserted into a different transgenic
bacterium. These can be maintained in their own cultures.
• The entire collection of these transgenic bacterial strains is called a gene library.
• Suppose you want to be tested to see if you carry BRCA1 (one of the major genes
involved in familial breast cancer).
• Your DNA can be collected. We can then take the BRCA1 gene from the human
gene library and see if it sticks to your DNA.
• The BRCA1 gene will stick if your DNA has complementary sequence to the
BRCA1 gene. If that is the case, you must carry the BRCA1 gene.
• In theory, we can do this with any other gene in our gene library (in practice,
not all of those genetic tests has yet been developed).
What can be done with transgenic organisms:
Genetically-Modified Crops and Livestock
What type of crops and livestock have been genetically modified?
Plants: Corn and Soy (Bt-Producing, Roundup Resistant)
Tomato (Slow ripening)
Strawberries (Frost-resistant)
Bananas (Hepatitis B antigen)
Animals: Cows (Bovine Growth Hormone)
Sheep (Fibrinogen production)
Chicken (Avian Flu Resistance)
Controversies
(http://www.ornl.gov/sci/techresources/Human_Genome/elsi/gmfood.shtml)
* Safety
* Access and Intellectual Property
* Ethics
* Labeling
* Society
What can be done with transgenic organisms:
Protein-Producing Bacteria
Genetically-modified bacteria are now grown in mass cultures to produce human
proteins. This is possible because bacteria read the genetic code in identical ways
to humans (and other organisms).
Human proteins produced by bacteria:
Insulin
Human Growth Hormone
Blood Clotting Factors
Erythropoietin
Interferon
What can be done with transgenic organisms:
Gene Therapy
Sick person’s cells
intentionally infected
with genetically
modified virus.
Good copy of human
gene from a donor
Person with a single
gene mutation that
causes a health
disorder
Harmless virus
genetically modified
to contain good copy
of human gene.
Viral DNA (including
good human gene)
integrates into sick
person’s chromosome.
Sick person’s cells begin
to produce functional
protein.
Sick person is CURED!