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CHAPTER 12
DNA Technology
PowerPoint® Lectures for
Campbell Essential Biology, Fourth Edition
– Eric Simon, Jane Reece, and Jean Dickey
Campbell Essential Biology with Physiology, Third Edition
– Eric Simon, Jane Reece, and Jean Dickey
Lectures by Chris C. Romero, updated by Edward J. Zalisko
© 2010 Pearson Education, Inc.
Stem Cells
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Adult stem
cells in
bone marrow
Blood cells
Nerve cells
Cultured
embryonic
stem cells
Heart muscle cells
Different culture
conditions
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Different types of
differentiated cells
Figure 11.15
Umbilical Cord Blood Banking
 Umbilical
 Can
cord blood
be collected at
birth
 Contains partially
differentiated stem
cells
 Limited use
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Figure 12.1
Recombinant DNA Techniques
•
•
Bacteria are the workhorses of modern biotechnology.
In the lab, biologists use bacterial plasmids (small, circular
DNA molecules) that are separate from the much larger
bacterial chromosome.
 Can
easily pick up foreign DNA
 Are taken up by bacterial cells; called transformation
 Act as vectors (DNA carriers that move genes from one cell to
another)

Recombinant DNA help biologists produce large quantities of a
desired protein.
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Bacterial
chromosome
Remnant of
bacterium
Colorized TEM
Plasmids
Figure 12.7
Cut both DNAs
with same
enzyme.
Gene of Other
interest genes
Gene of interest
Bacterial cell
DNA fragments
from cell
Isolate
DNA.
Mix the DNAs and
join them together.
Cell containing
the gene of interest
Isolate
plasmids.
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
Plasmid
DNA
Bacterial clone
Recombinant bacteria
Clone the bacteria.
Find the clone with
the gene of interest.
Some uses
of genes
Gene for pest
resistance
Some uses
of proteins
Protein for
dissolving
clots
Gene for
toxic-cleanup
bacteria
Genes may be
inserted into
other organisms.
The gene and protein
of interest are isolated
from the bacteria.
Harvested
proteins may be
used directly.
Protein for
“stone-washing”
jeans
Figure 12.8-8
A Closer Look: Cutting and Pasting DNA
with Restriction Enzymes
 Recombinant
DNA is produced by combining two
ingredients:
A
bacterial plasmid
 The gene of interest
 To
combine these ingredients, a piece of DNA must be
spliced into a plasmid.
 This splicing process can be accomplished by:
 Using
restriction enzymes, which cut DNA at specific nucleotide
sequences
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Restriction enzymes

Bacterial enzymes that recognize and cut DNA at
specific sequences
 What

is there use naturally in bacteria?
Are very specific
 Usually
recognize sequences 4-8 nucleotides long
 Sequences recognized are palindromes
 Example: EcoR1 recognizes GAATTC and cuts always
between the G and A
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• Producing pieces of DNA called restriction fragments with
“sticky ends” important for joining DNA from different sources
– DNA ligase connects the DNA pieces into continuous
strands by forming bonds between adjacent nucleotides.
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Recognition sequence
for a restriction enzyme
DNA
A restriction enzyme cuts
the DNA into fragments.
Restriction
enzyme
Figure 12.9-1
Recognition sequence
for a restriction enzyme
DNA
A restriction enzyme cuts
the DNA into fragments.
Restriction
enzyme
A DNA fragment is added
from another source.
Figure 12.9-2
Recognition sequence
for a restriction enzyme
DNA
A restriction enzyme cuts
the DNA into fragments.
Restriction
enzyme
A DNA fragment is added
from another source.
Fragments stick together by
base pairing.
Figure 12.9-3
Recognition sequence
for a restriction enzyme
DNA
A restriction enzyme cuts
the DNA into fragments.
Restriction
enzyme
A DNA fragment is added
from another source.
Fragments stick together by
base pairing.
DNA ligase joins the
fragments into strands.
DNA
ligase
Recombinant DNA molecule
Figure 12.9-4
Genetic engineering vocab
 Recombinant
DNA- nucleotide sequences from two
different sources to form a single DNA molecule.
 genetic engineering, the direct manipulation of DNA for
practical purposes.
 Biotechnology – use of organisms or their components to
make useful products
 Transgenic organism – contains a gene from another
organism, typically a different species
 Genetically modified organisms (GMOs)- organisms that
have acquired one or more genes by artificial means.
© 2010 Pearson Education, Inc.
Cut both DNAs
with same
enzyme.
Gene of Other
interest genes
Gene of interest
Bacterial cell
DNA fragments
from cell
Isolate
DNA.
Mix the DNAs and
join them together.
Cell containing
the gene of interest
Isolate
plasmids.
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
Plasmid
DNA
Bacterial clone
Recombinant bacteria
Clone the bacteria.
Find the clone with
the gene of interest.
Some uses
of genes
Gene for pest
resistance
Some uses
of proteins
Protein for
dissolving
clots
Gene for
toxic-cleanup
bacteria
Genes may be
inserted into
other organisms.
The gene and protein
of interest are isolated
from the bacteria.
Harvested
proteins may be
used directly.
Protein for
“stone-washing”
jeans
Figure 12.8-8
Warm up December 17

How do these cats show
examples of genetic
engineering, transgenic
organisms and
recombinant DNA?
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Gene for human growth
hormone
DNA recombination
Human Cell
Sticky ends
DNA insertion
Bacterial Cell
Bacterial
chromosome
Bacterial cell for containing gene
for human growth hormone
Plasmid
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Making Humulin - 1st engineered product
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Genetically Modified (GM) Foods
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 “Golden
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rice”
DNA PROFILING AND FORENSIC SCIENCE
 DNA
profiling (fingerprinting):
 Used
to determine if two samples of genetic material are from
same person
 Scientific crime scene analysis
 To
produce a DNA profile
 Scientists

compare genetic markers
Sequences in the genome that vary from person to person.
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Investigating Murder, Paternity, and Ancient DNA
 DNA
profiling can be used to:
 Test
the guilt of suspected criminals
 Identify tissue samples of victims
 Resolve paternity cases
 Identify contraband animal products
 Trace the evolutionary history of organisms
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Crime scene
Suspect 1
Suspect 2
DNA isolated
DNA amplified
DNA compared
Figure 12.13-3
DNA Profiling Techniques
The Polymerase Chain Reaction (PCR)
 The
polymerase chain reaction
(PCR):
 Is
a technique to copy quickly and
precisely any segment of DNA
 Can
generate enough DNA, from
even minute amounts of blood or
other tissue, to allow DNA
profiling
How do you test if two samples of DNA come from the
same person?
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Gel Electrophoresis
 Compares
the lengths of varied DNA fragments
 Uses
gel electrophoresis, a method for sorting
macromolecules—usually proteins or nucleic acids—primarily by
their


Electrical charge
Size
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Mixture of DNA
fragments of
different sizes
Band of
longest
(slowest)
fragments
Power
source
Gel
Completed gel
Band of
shortest
(fastest)
fragments
•Shorter fragments travel through the gel faster than longer fragments
•Fragments travel to positive end because phosphates in DNA are
negatively charged
•2 ways to analyze DNA in gel electrophoresis
Short Tandem Repeat (STR) Analysis
 Short
Tandem Repeats (STR’s)
 Short
repetitions (usually 4 nucleotides)
 Number of repeats can vary from person to person
 Used in DNA profiling/criminal investigations

FBI uses 13 repetitive sites on our DNA
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STR site 1
AGAT
STR site 2
GATA
Crime scene DNA
Different numbers of
short tandem repeats
Same number of
short tandem repeats
Suspect’s DNA
AGAT
GATA
Figure 12.16
Amplified
crime scene
DNA
Amplified
suspect’s
DNA
Longer
fragments
Shorter
fragments
Figure 12.18
RFLP Analysis

Before placed in the gel, DNA is mixed and cut by
restriction enzymes
 Individuals
have unique restriction sites so DNA fragment lengths
may vary
 Used often to compare different gene alleles
 Basis of some genetic and paternity tests
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RFLP
EcoRI
GAATTC
CTTAAG
ATGCTTAAGGCGTACACTGAATTCTAGTACCTA
TACGAATTCCGCATGTGACTTAAGATCATGGAT
ATGCTTAAGGCGTACACTG
TACGAATTCCGCATGTGACTTAA
AATTCTAGTACCTA
GATCATGGAT
Region cut into 2 fragments by EcoRI
ATGCTTAAGGCGTACACTGGATTTCTAGTACCTA
TACGAATTCCGCATGTGACCTTAAGATCATGGT T
ATGCTTAAGGCGTACACTGGATTTCTAGTACCTA
TACGAATTCCGCATGTGACCTTAAGATCATGGT T
Region not cut by EcoRI due to base substitution at restriction site.
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Restriction enzymes added
Suspect’s
DNA
Crime scene
DNA
Fragment w
Cut
Fragment z
Fragment x
Cut
Cut
Fragment y
Fragment y
Crime scene
DNA
Longer
fragments
Suspect’s
DNA
z
x
Shorter
fragments
w
y
y
Figure 12.19
Lane 2 is mom, lane 5 is son
so…who’s the daddy? 3 or 4?
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For the test
1.
Cloning
1.
2.
3.
2.
Stem cells
1.
2.
3.
4.
3.
What is it?
Reproductive vs. therapeutic cloning
How to do reproductive cloning (how did you clone mimi the mouse)
What are they?
Types of stem cells and their potential
Where do we find different types of stem cells such as adult, embryonic
What are IPS stem cells and why are they important?
Recombinant DNA
1.
2.
3.
What is it?
How do we make a recombinant plasmid?
What is it used for?
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For the test
PCR/gel electrophoresis
4.
4.
5.
6.
What are they used for?
How is a gel electrophoresis run?
How is a gel electrophoresis read?
Human genome project/Gene therapy
5.
4.
5.
What are they?
For what do they hope to use these?
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Table 12.1
The Human Genome Project
 Begun
in 1990, the Human Genome Project was a massive
scientific endeavor:
 To
determine the nucleotide sequence of all the DNA in the human
genome and
 To identify the location and sequence of every gene
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HUMAN GENE THERAPY
 Human
 Is
gene therapy:
a recombinant DNA procedure
 Seeks to treat disease by altering the genes of the afflicted
person
 Often replaces or supplements the mutant version of a gene with
a properly functioning one
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Normal human
gene isolated
and cloned
Healthy person
Figure 12.24-1
Normal human
gene isolated
and cloned
Harmless
virus (vector)
Normal human
gene inserted
into virus
Virus containing
normal human gene
Healthy person
Figure 12.24-2
Normal human
gene isolated
and cloned
Harmless
virus (vector)
Normal human
gene inserted
into virus
Virus containing
normal human gene
Bone
marrow
Healthy person
Virus injected
into patient with
abnormal gene
Bone of person
with disease
Figure 12.24-3
SCID – severe combined immune
deficiency
 SCID
is a fatal inherited disease caused by a single
defective gene that prevents the development of the
immune system.
 SCID patients quickly die unless treated with:
A
bone marrow transplant or
 Gene therapy
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SCID and gene therapy
 Since
the year 2000, gene therapy has:
 Cured
22 children with inborn SCID but
 Unfortunately, caused four of the patients to develop
leukemia, killing one of these children
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