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CHAPTER 15
Studying and Manipulating Genomes
How are Genetic changes produced?
 Natural processes

Mutation, variation
 Humans have been changing the genetics of
many species for thousands of years

Artificial selection of plants and animals
 Humans changing genetics by combining
genes between species

Genetic Engineering
Purpose of Genetic Engineering
 Cure hereditary
diseases
 Engineer livestock to


Provide more nutrients
Make vaccines, vitamins
 Engineer crops to



Produce pesticides
Increase productivity
Better nutrients
•
Basic research

Investigate basic genetic
processes
Reconstruct life’s
evolutionary history
Devise counterattacks
against rapidly mutating
pathogens


First Recombinant DNA
 Paul Berg and associates were first to make
recombinant DNA in 1972
 Fused fragments of DNA from one species into the
genetic material from another
 Allowed them to isolate and replicate subsets of DNA
from any organism
Tools for Constructing Recombinant
DNA
Restriction Enzymes
 Hamilton Smith was studying Haemophilus
influenzae discovered bacteria have an enzyme that
chops up viral DNA.
 Will cut at a specific site on a specific sequence of
DNA. The section has to be a palindrome!

Ex. GAATTC is cut by EcoRI
The same enzyme is used for both the DNA fragments so
the overhanging ends are identical and complementary

Ligase used to glue them together

Making Recombinant DNA
Tools for Constructing Recombinant
DNA
Plasmids as Vectors
 Small circular DNA pieces,
 can be used to transfer foreign DNA to bacteria
 Why: So that the gene can be expressed and
studied
Can also be inserted into eukaryotic cells
directly Ex. Humans, plants etc.
Making Recombinant DNA
Recombine DNA to be studied with a Plasmid
DNA. Transform bacteria with this plasmid. Can
be studied and manipulated.
Tools for Constructing Recombinant DNA
PCR – Polymerase Chain reaction
Many rounds of DNA replication in a test tube to
generate thousands of copies of a DNA segment.
Used for

forensics

Insertion and cloning

sequencing of DNA
Polymerase Chain Reaction
Polymerase Chain Reaction
Polymerase Chain Reaction
Polymerase Chain Reaction
DNA replication in a tube!
1. Sequence to be copied is heated to break H
bonds
2. Primers are added and bind to ends of single
strands
3. Taq DNA polymerase uses free nucleotides to
create complementary strands
4. Repeat Steps 1-3
5. Doubles number of copies of DNA at each
round
Tools for Constructing Recombinant
DNA
Gel Electrophoresis
Technique by which
molecules of different sizes
are separated and analyzed
 Separated by electric
charge - DNA is
negatively charged and
will migrate towards
positive electric field
 Rate of migration
depends on size of
molecule
+
Tools for Constructing Recombinant
DNA
Sequencing DNA
• Fluorescent labeled and
fragmented DNA placed on gel
• Fragments move off gel in
size order; pass through laser
beam
• Color each fragment
fluoresces is recorded on
printout
Uses of genetic engineering: The Human
Genome Project
Goal – Use these tools to map the entire human
genome
 Initially thought by many to be a waste of
resources
 Process accelerated when Craig Venter used bits
of cDNAs as hooks to find genes
 Sequencing was completed ahead of schedule in
early 2001
Human Genome
Uses of genetic engineeringDNA
Fingerprinting
• Unique array of DNA
fragments
• Inherited from
parents
• Even full siblings can
be distinguished from
one another by this
technique
Analyzing DNA Fingerprints
• DNA is stained or made
visible by use of a
radioactive probe
• Pattern of bands is used
to:
–
Identify or rule out
criminal suspects
–
Determine paternity