Genetic Engineering
Chapter 10
What we are hoping to accomplish
•
Define genetic engineering.
•
Explain the principles of genetic engineering.
•
Explain and use the tools of genetic engineering.
•
Show and describe the practical uses of genetic
engineering.
•
Discuss the various steps in PCR and
transformation.
Biotechnology
-
Genetic Engineering
•
The application of biological and physical
principles to alter the genes of organisms to help
solve human problems
▫
Improving crops and their yield
▫
Producing insulin in bacteria
▫
Genetically modified vaccines
▫
Improving the freshness of fruits and vegetables
Biotechnology
Historical
Aspects
Plant breeding
Animal Breeding
Wine making
Dairy products
Bread
Antibiotics
Herbal medicine
Business and
Industry
Biomedical
Agriculture
Forensics
Industrial
production
Conservation
Wildlife
management
Multidisciplinary
Research
Biochemistry
Molecular genetics
Biophysics
Immunology
Cell culture
Fermentation
Bacteriology
Bioengineering
Genetic medicine
Conservation
Evolutionary
biology
Social and Ethical
Implications
Genetic privacy
Ownership of info.
Screening
Engineered food
Insurance
Treatment
DNA profiling
Patenting of life
What does genetic engineering mean?
•
It means to isolate a gene from the DNA of one
organism and transfer the gene into the DNA of
another.
•
We can do this because DNA is DNA in all
organisms. It is made out of the same 4 nitrogen
bases. The only difference is the sequence.
•
From there, DNA codes for RNA and proteins.
Recombinant DNA
•
A molecule of DNA made from pieces of DNA
from separate organisms.
Restriction Enzyme
•
An enzyme that is used to “cut” DNA at certain
places along
BOTH
sides of the DNA.
•
Most commonly used Restriction Enzymes
▫
BamHI
-
G/GATTC
▫
HindIII
-
A/AGCTT
▫
HpaII
-
C/CGG
▫
EcoR1
-
G/AATTC
Plasmids
•
Circular piece of DNA found in bacteria.
▫
Replicates separate chromosome DNA
▫
Contains very few genes
▫
Useful because they have antibiotic resistance
genes on them.
▫
Typically used as
VECTORS
to transfer DNA
from one organism to another.
Polymerase Chain Reaction (PCR)
•
A way to make an unlimited number of copies of
one gene.
•
How? DNA Polymerase is used to replicate the
same DNA segment over and over.
•
Why?
▫
Increasing the number (Crime Scenes)
▫
Sequencing (Prenatal diagnosis)
▫
Cloning (Woolly Mammoth)
3 Steps to PCR
1.
Denaturing
-
Stands of DNA separate by
heating
2.
Annealing
-
Reaction cooled, primers bind
3.
Elongation
-
Reaction heated again, DNA
polymerase replicates the rest of the DNA from
primer.
•
Repeat steps 1
-
3 about 20
-
30x
DNA Fingerprinting
•
DNA is sliced into fragments called RFLPs.
•
DNA is then separated by sequence length by gel
electrophoresis.
•
The result is for an individual different lengths
of DNA that were cut by restriction enzymes that
varies among alleles and individuals.
Gel Electrophoresis
How the Gel Works?
•
DNA is negative
•
When DNA is placed next to a negative charge it
will want to move towards the positive charge.
•
The heavier strands will not travel as far as the
lighter strands.
DNA fingerprinting in a crime scene
Who did it? How Do We Know?
Crime Scene
Suspect 1
Suspect 2
Suspect 3
Suspect 4
Use in Crime Scenes
•
3 different Fingerprints are run
•
1/10,000 chance of matching an individual
•
Two fingerprints will only match 1 out of 10,000
X 10,000 times
•
Three matching prints would then be 1 in one
trillion
The DNA
•
Magnified using PCR
•
From blood, bone, flesh, hair follicle, semen,
saliva, any source of DNA
•
Analysis does not compare genes rather the
information between genes where a great
amount of variation exists
▫
RFLP analysis
Restriction Fragment Length
Polymorphism (RFLP) Analysis
•
Identifies locations and numbers of restriction
sites of DNA.
Human Genome Project
•
Purpose: Sequence the entire human genome.
•
Why: detect, treat, and prevent genetic diseases.
•
Completed in 2003
•
Some of their findings for human DNA
▫
Approximately 20,000
-
25,000 genes
(23,000)
▫
3 billion base pairs make up the DNA.
▫
Only 1.5% is actually coding for protein
•
Still left to do? Figure out how it all fits together.
Typical Transformation Experiment
1.
A target gene is isolated and removed by a
restriction enzyme
.
2.
Another restriction enzyme will cut the DNA of
a
vector
(
plasmids
) and insert our target
gene.
3.
Transfer the vector to the organism we want to
modify.
4.
Transformation
-
the gene becomes one with
its host and replicates producing clone copies.
5.
Screen the cells and select modified organism
by identifying our gene of interest.
Why do we care?
Genetically Engineered Drugs
Genetically Engineered Vaccines
•
Humulin
-
Diabetes
•
HGH
-
Dwarfism
•
Factor VIII, Factor IX
-
Hemophilia
•
t
-
PA
–
Heart attack, clot
buster
•
Lactoferrin
–
Milk protein
•
Herpes II
•
Hepatitis B
•
Influenza
Why do we care?
Cures for Genetic Diseases
Forensic Science
•
Cystic fibrosis
•
Genetic testing
•
Therapy
•
Karyotyping
•
DNA fingerprints
▫
Crime
▫
Paternity
▫
Predisposed to disease
This may involve using stem
cells, by taking somatic cells
(like skin cells) and
dedifferentiating them down
to
pluripotent
stem cells.
Why do we care?
Plants
Animals
•
Ti plasmid causes plant
tumors
•
Glyphosate
resistant crops
(round
-
up ready)
•
Pest resistance
•
Improve taste and nutrition
•
Vaccines grown in plants
•
BST
•
Lactoferrin
•
Cloning
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