Download Chapter 8: Gene Expression, Mutation, Cloning

Chapter 8
 Genetically Modified Organisms
 Gene Expression, Mutation, and Cloning
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Chapter 8 Section 1
Protein Synthesis and Gene Expression
Part A: Transcription
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Genetic Engineering
Genetic Engineering
 Alteration of hereditary traits by molecular
biological techniques
 One or more genes may be modified
 Genes may be moved from one organism to
another
 Move a gene that produces a desired
protein from one organism to another
 Alter regulation of gene expression
 Change the amount of protein that a gene
produces
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Genetic Engineering
Genetic Engineering Controversy
 Benefits:
 Potential to wipe out hunger by make crops
more productive
 Vaccinate children by eating foods
 Reduce heart problems by added omega-3
fatty acids to animals like pigs
 Concerns:
 Is it ethical?
 Is it safe for use in food sources?
 Will it be used in humans?
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8.1 Protein Synthesis and Expression
History of Genetic Engineering
 In the early 1980s, genetic engineers at
Monsanto® Company began producing
recombinant bovine growth hormone
(rBGH)
 Made by genetically engineered bacteria
 The bacteria were given DNA that carries
instructions for making BGH
 Giving growth hormone to cows increase
body size and milk production
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8.1 Protein Synthesis and Expression
How do genes make proteins?
Protein synthesis
 the process of using instructions carried on
a gene to create proteins.
 Gene – a sequence of DNA that encodes a
protein
 Protein – a large molecule composed of
amino acids
 Several steps are involved and require
both DNA and RNA.
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8.1 Protein Synthesis and Expression
Structure of DNA & RNA
DNA
 Double-stranded
 Each nucleotide
composed of
deoxyribose,
phosphate, and
nitrogenous base
 4 bases: adenine,
thymine, guanine,
cytosine
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8.1 Protein Synthesis and Expression
Structure of DNA & RNA
RNA
 Single-stranded
 Nucleotides
comprised of
ribose,
phosphate, and
nitrogenous base
 4 bases: A, C, G,
and Uracil
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8.1 Protein Synthesis and Expression:
From Gene to Protein
 The flow of genetic information in a cell is
DNA  RNA  protein
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Figure 8.2
8.1 Protein Synthesis and Expression:
From Gene to Protein
 There are 2 steps in going from gene to protein
 Transcription (DNA  RNA)
 Translation (RNA  Protein)
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8.1 Protein Synthesis and Expression:
Transcription
 Transcription occurs in the nucleus.
 RNA polymerase binds to the promoter region
of the gene.
 RNA polymerase zips down the length of gene,
matching RNA nucleotides with complementary
DNA nucleotides
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8.1 Protein Synthesis and Expression:
Transcription
 The product of transcription is
messenger RNA (mRNA).
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8.1 Protein Synthesis and Expression:
Transcription
PLAY
Animation—Transcription
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Chapter 8 Section 1
Protein Synthesis and Gene Expression
End Part A: Transcription
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Chapter 8 Section 1
Protein Synthesis and Gene Expression
Part B: Translation and the Genetic Code
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8.1 Protein Synthesis and Expression:
Translation
Translation
 Uses mRNA template to make protein
 Translation occurs in the cytoplasm
 Translation requires:





mRNA
amino acids
ATP
Ribosomes
Transfer RNA (tRNA)
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8.1 Protein Synthesis and Expression:
Translation
Ribosomes
 The ribosome is composed of
rRNA and comprises a small
and a large subunit.
 When subunits come together,
mRNA can be threaded
between them to make protein
 Also requires transfer RNA
(tRNA) to bring correct amino
acids
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Large
subunit
Small
subunit
Figure 8.4
8.1 Protein Synthesis and Expression:
Translation
Transfer RNA (tRNA )
 Each tRNA carries
one specific amino
acids and matches
its anticodon with
codons on mRNA
 So what are
codons and
anticodons??
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Amino
acid
Binding site for
amino acid
Region of internal
complementarity
tRNA
Anticodon
mRNA
Codon
Figure 8.5
8.1 Protein Synthesis and Expression:
Translation
As ribosome moves along mRNA, small
sequences of nucleic acids are exposed
 Codon – a three letter genetic code of
nucleic acids in mRNA that indicate a
specific amino acid
 Anticodon – a complementary three
nucleic acid sequence on tRNA that
matches codon
 >By using anticodons to match codons,
tRNA can bring the correct amino acid
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Figure 8.5
8.1 Protein Synthesis and Expression:
Genetic Code
The genetic code
 The use of nucleic acid codons to specify
amino acid sequence in proteins
 A codon is comprised of three nucleotides = 64
possible combinations (43 combinations)
 61 codons code for ~20 amino acids
 Redundancy – may be more than 1 code
per amino acid
 3 others are stop codons, which end protein
synthesis
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8.1 Protein Synthesis and Expression:
Genetic Code
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Table 8.1
8.1 Protein Synthesis and Expression:
Translation
A protein is put together one amino acid at a
time.
1. The ribosome attaches to the mRNA at the
promoter region.
2. Ribosome facilitates the docking of tRNA
anticodons to mRNA codons.
3. When two tRNAs are adjacent, a bond is
formed between their amino acids.
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8.1 Protein Synthesis and Expression:
Translation
Translation Step by Step
1 Amino acids and tRNAs float
freely in the cytoplasm.
Amino acid
2 Enzymes facilitate the binding of a specific tRNA to its
appropriate amino acid.
tRNA
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Figure 8.7
8.1 Protein Synthesis and Expression:
Translation
3 A tRNA will dock if the
complementary RNA codon
is present on the ribosome.
4 The amino acids join together
to form a polypeptide.
Amino acid chain (polypeptide)
ala phe ile
Stop codon
AAA UAU
GCCUUUAUA
Ribosome
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Figure 8.7
8.1 Protein Synthesis and Expression:
Translation
Amino acid chain (polypeptide)
ala phe ile
Stop codon
AAA UAU
GCCUUUAUA
5 The ribosome
moves on to
the next
codon to
receive the
next tRNA.
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Ribosome
6 When the ribosome reaches the
stop codon, no tRNA can basepair with the codon on the mRNA.
RNA and the newly synthesized
protein are released.
Figure 8.7
8.1 Protein Synthesis and Expression:
Translation
7 The chain of amino
acids folds, and the
protein is ready to
perform its job.
AGC STOP
CUCUCGUAA
Protein
(such as BGH)
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8 The subunits of the ribosome
separate but can reassemble
and begin translation of
another mRNA.
Figure 8.7
8.1 Protein Synthesis and Expression:
Translation
PLAY
Animation—Translation
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Chapter 8 Section 1
Protein Synthesis and Gene Expression
END Part B: Translation and the Genetic Code
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Chapter 8 Section 1
Protein Synthesis and Gene Expression
Part C: Types of Mutations and Regulating
Gene Expression
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8.1 Protein Synthesis and Expression:
Mutations
Mutations = Changes in genetic sequence
 Changes in genetic sequence might affect
the order of amino acids in a protein.
 Protein function is dependent on the precise
order of amino acids
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8.1 Protein Synthesis and Expression:
Mutation
Possible outcomes of mutation:
1 - no change in protein (neutral mutation)
2 - non-functional protein
3 - different protein
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Figure 8.8
8.1 Protein Synthesis and Expression:
Mutation
Types of Mutations
1. Base-substitution mutation – simple
substitution of one base for another
2. Frameshift mutation – addition or
deletion of a base, which changes the
reading frame
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8.1 Protein Synthesis and Expression
Examples of Mutations
• Neutral Mutation
• Frameshift Mutation
- adding or deleting a
nucleic acid
- shifts entire
sequence
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Figure 8.9
8.1 Protein Synthesis and Expression:
Overview of Gene Expression
 Each cell in your body (except sperm and
egg cells) has the same DNA.
 But each cell only expresses a small
percentage of genes.
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8.1 Protein Synthesis and Expression: An
Overview of Gene Expression
 Regulating gene expression
 Turning a gene or a set of genes on or off
 EXP: Nerves and muscles have the same
suite of genes, but express different genes.
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Figure 8.11
8.1 Protein Synthesis and Expression:
 Regulating gene expression
1. Repression of transcription
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Figure 8.11
8.1 Protein Synthesis and Expression:
 Regulating gene expression
2. Activation of transcription
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Figure 8.11
Chapter 8 Section 1
Protein Synthesis and Gene Expression
Part C: Types of Mutations and Regulating
Gene Expression
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Chapter 8 Section 2
Producing Recombinant Proteins
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8.2 Producing Recombinant Proteins:
Cloning a Gene Using Bacteria
Producing Recombinant Proteins
 BGH is a protein, and is coded by a
specific gene.
 Transfer of BGH gene to bacteria allows for
growth under ideal conditions.
 Bacteria can serve as “factories” for
production of BGH.
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8.2 Producing Recombinant Proteins:
Cloning a Gene Using Bacteria
Two Important molecules in cloning
1. Restriction enzymes
2. Plasmids
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8.2 Producing Recombinant Proteins:
Cloning a Gene Using Bacteria
Restriction enzymes
 Used by bacteria as a form of defense.
 Restriction enzymes cut DNA at specific
sequences.
 Leave “sticky ends”
 They are important in biotechnology because
they allow scientists to make precise cuts in
DNA.
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8.2 Producing Recombinant Proteins:
Cloning a Gene Using Bacteria
Plasmid
 Small, circular piece of bacterial DNA that exists
separate from the bacterial chromosome.
 Plasmids are important because they can act as
a ferry to carry a gene into a cell.
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8.2 Producing Recombinant Proteins:
Steps in Cloning a Gene Using Bacteria
 Step 1. Remove the gene from the cow
chromosome using restriction enzymes
1 BGH gene is cut from the cow chromosome
using restriction enzymes that leave “sticky
ends” with specific base sequences.
Cow cell
BGH
gene
DNA
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Figure 8.13
8.2 Producing Recombinant Proteins:
Cloning a Gene Using Bacteria
 Step 2. Insert the BGH gene into the
bacterial plasmid
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Figure 8.13
8.2 Producing Recombinant Proteins:
Cloning a Gene Using Bacteria
 Recombinant – Indicates material that
has been genetically engineered. A gene
that has been removed from its original
genome and combined with another.
 After step 2, the BGH is now referred to as
recombinant BGH or rBGH.
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8.2 Producing Recombinant Proteins:
Cloning a Gene Using Bacteria
 Step 3. Insert the recombinant
plasmid into a bacterial cell
Recombinant
plasmid
3 The recombinant plasmid is
reinserted into a bacterial cell.
The plasmids and the bacterial cells
replicate, making millions of copies of
the rBGH gene.
rBGH
proteins
The rBGH genes produce large
quantities of rBGH proteins that are
harvested, purified, and injected into
cows to increase milk production.
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Figure 8.13
8.2 Producing Recombinant Proteins
PLAY
Animation—Producing Bovine Growth Hormone
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8.2 Producing Recombinant Proteins
Use of rBGH in Agriculture
 About 1/3 of cows in the US are injected
with rBGH.
 rBGH increases milk volume from cows by
about 20%.
 Increase profits for both ranchers and
industry
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8.2 Producing Recombinant Proteins
Controversy over Use of rBGH
 Controversy over safety to humans
 USDA and Monsanto argue that milk from
rBGH-treated cows is indistinguishable from
non-treated.
 Activists disagree.
Welfare of cows?
Europe and Canada banned
rBGH over concerns on the
health of cows.
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8.2 Producing Recombinant Proteins:
Cloning a Gene Using Bacteria
The same principles apply to other proteins
 Clotting proteins for hemophiliacs are
produced using similar methods.
 Insulin for diabetics is also produced in this
way.
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END Chapter 8 Section 2
Producing Recombinant Proteins
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Chapter 8 Section 3
Genetically Modified Foods
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8.3 Genetically Modified Foods
Artificial Selection
 All agricultural products
are the result of genetic
modification through
selective breeding.
 Artificial selection
does not move genes
from one organism to
another, but does
drastically change the
characteristics of a
population.
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Figure 8.13
8.3 Genetically Modified Foods:
Why Genetically Modify Crop Plants?
 Increase shelf life, yield, or
nutritional value
 Exp: Golden rice has been
genetically engineered to
produce beta-carotene,
which increases the rice’s
nutritional yield.
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Figure 8.16
8.3 Genetically Modified Foods: Modifying
Plants with the Ti Plasmid and Gene Gun
Modifying Plants Directly
 (Different than rBGH, which was a protein
injected into cows)
 In order to do this, the target gene must be
inserted into the plant cell.
 Two methods can inject genes into plants:
1. Ti plasmid
2. Gene gun
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8.3 Genetically Modified Foods: Modifying
Plants with the Ti Plasmid and Gene Gun
Modifying Plants wiith the Ti plasmid
 To modify the plant, must get gene across
cell wall
 The bacterium Agrobacterium tumefaciens
does this naturally, creating ‘galls’
• The bacterium can do this
by using the Ti plasmid
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8.3 Genetically Modified Foods:
Modifying Plants wiith the Ti plasmid
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Figure 8.17b
8.3 Genetically Modified Foods: Modifying
Plants with the Ti Plasmid and Gene Gun
 The Gene Gun
Microscopic
particles coated
with gene of
interest are
“shot” into
plant cells.
Gun
Shock waves
“Bullet”
Plant cells
in culture
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Figure 8.18
8.3 Genetically Modified Foods:
Transgenic organism
 the result is the incorporation of a gene
from one organism to the genome of
another.
 Also referred to as a Genetically Modified
Organism (GMO).
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8.3 Genetically Modified Foods: Effect of GM
Crops and the Environment
Benefits
 Crops can be engineered for resistance to
pests, thus farmers can spray fewer
chemicals.
Concerns
 GM crops may actually lead to increased
use of pesticides and herbicides.
 EXP: Roundup-Ready plants
 GM crop plants may transfer genes to wild
relatives.
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End Chapter 8 Section 3
Genetically Modified Foods
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Chapter 8 Section 4
Genetically Modified Humans
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8.4 Genetically Modified Humans: Stem Cells
Genetically Modified Humans
 Stem cells:
 undifferentiated cells
 Totipotent = capable of
growing into many
different kinds of cells
and tissues.
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Figure 8.20
8.4 Genetically Modified Humans: Stem Cells
Use of Stem cells
 Stem cells can be collected, with consent,
from left over embryos from artificial
fertilization
 Cells can be grown ‘in vitro’ in the lab
 Can be ‘directed’ to develop into many
different kinds of tissues
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8.4 Genetically Modified Humans: Stem Cells
Therapeutic Cloning
 Stems cells might be used to treat
degenerative diseases
 Alzheimer’s or Parkinson’s, multiple
sclerosis, or liver, lung, or heart disease.
 Stem cells could also be used to grow
specific tissues
 to treat burns, heart attack damage,
replacement cartilage in joints, or spinal cord
injuries.
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8.4 Genetically Modified Humans:
Human Genome Project
 international effort to map the sequence of
the entire human genome (~20,000 –
25,000 genes).
 A genome is all of the genes in an organism
 For comparative purposes, genomes of other
model organisms (E. coli, yeast, fruit flies,
mice) were also mapped.
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8.4 Genetically Modified Humans:
Gene Therapy
= replacement of defective genes with
functional genes
Two approaches:
1. Germ line gene therapy
2. Somatic cell gene therapy
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8.4 Genetically Modified Humans: Gene
Therapy
Germ line gene therapy
 Embryonic treatment
 Embryo supplied with a functional version
of the defective gene.
 Embryo + cells produced by cell division
have a functional version of gene.
 Including their children
 > would prevent genetic diseases
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8.4 Genetically Modified Humans: Gene
Therapy
Somatic cell gene therapy
 fix or replace the defective protein only in
specific cells.
 Performed on adult body cells
 Put copy of good gene into cells in lab,
multiply cells, then introduce to affected
person
 Already used as a treatment of SCID (severe
combined immunodeficiency)
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8.4 Genetically Modified Humans: Gene
Therapy
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8.4 Genetically Modified Humans:
Cloning Humans
 Cloning = making a genetically identical
organism
 Cloning occurs naturally whenever identical
twins are produced.
 Cloning of offspring from adults has
already been done with cattle, goats, mice,
cats, pigs, and sheep.
 Cloning is achieved through the process of
NUCLEAR TRANSFER.
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8.4 Genetically Modified Humans:
Cloning Dolly the Sheep
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8.4 Genetically Modified Humans: Cloning
Humans
 Genetic engineering is controversial.
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Table 8.2
END Chapter 8 Section 4
Genetically Modified Humans
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END Chapter 8
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