Transcript DNA Technology and Genomes

Genetic Engineering
and Biotechnology
IB Biology HL I
Spring 2014
Mrs. Peters
What do we already know?
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What does DNA contain?
What is the function of a gene?
What do you need to make a protein?
How are prokaryotes and eukaryotes
different?
What is Genetic Engineering?
Genetic Engineering: the direct
manipulation of DNA for practical
purposes
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used in Forensic Science, agriculture,
medicine and food technology
Biotechnology: the manipulation (through
genetic engineering) of living organisms or
their components to produce useful
usually commercial products
Important Terms
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Gene Cloning: making multiple identical
copies of a gene (specific pieces of DNA)
Clone: a group of genetically identical
organisms or a group of cells derived from a
single parent cell.
Plasmid: circular DNA found in bacteria, not
part of the nucleoid region
Restriction Enzymes: enzymes that protect
bacteria by cutting up foreign DNA from
invaders
DNA ligase: enzyme used to seal DNA
strands together
Important Terms
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Restriction site: the same sequence of 4 to
8 nucleotides, usually symmetrical, where
restriction enzymes cut DNA
Restriction Fragment: the piece of DNA
that is cut out of a DNA strand by restriction
enzymes.
Sticky ends: short single stranded
sequences on both sides of a restriction
fragment of DNA
Recombinant DNA: a “new” DNA strand
which contains the original DNA + a
restriction fragment. (recombined DNA)
Restriction Enzymes
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Enzymes that cut DNA molecules at
specific locations
Discovered in late 1960’s
Found in Prokaryotes, used naturally to
protect bacteria from invading DNA from
other organisms
Used in science to make DNA fragments
and recombinant DNA
Making Recombinant DNA
1. Restriction enzymes
recognize a specific DNA
sequence
2. Restriction enzyme cuts
DNA, producing sticky ends
3. DNA fragment from other
source is added; fragments
stick together by base
pairing
4. DNA ligase seals the
strands, resulting in
recombinant DNA molecule
Time to Practice!
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RFLP Activity-DNA Scissors
DNA Cloning
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Process of making
multiple identical
copies of a gene
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aka gene cloning
DNA technology uses
bacteria plasmids to
clone genes
DNA and Gene Cloning Process
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2.
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Isolate plasmid DNA
from bacteria and DNA
gene from other
organism
Using a restriction
enzyme, cut the plasmid
and the gene from the
other DNA strand
Gene is inserted into
plasmid and sealed with
DNA Ligase, forming
recombinant DNA
DNA and Gene Cloning Process
4. Plasmid put into bacterial
cell
5. Cell grown in culture,
forming clones (lots of
copies)
6. Desired gene is
identified by products
formed (protein or
characteristic)
Applications of Cloning
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Copies of Genes:
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For pest resistance in
plants
For basic research on
genes
To alter bacteria for
cleaning toxic waste
Copies of proteins:
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Human growth hormone
treatments
For basic research on
proteins
Dissolving blood clots in
heart attack therapy
Time to Practice
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Plasmid Simulation
DNA Profiling
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DNA profiling is the process of matching
DNA from a collected sample to a known
individual.
DNA samples come from hair, skin, blood
and other body fluids
DNA profiling is used in forensic science to
establish the possibility of guilt or prove a
suspect innocent; also used in paternity
testing
What happens on CSI and NCIS
PCR
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Polymerase Chain
Reaction
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a technique in which a
specific piece of DNA
is copied quickly
without the use of cells
Used for DNA profiling
in forensic science and
paternity testing
PCR: What is the machine doing?
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PCR Process:
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DNA is incubated in a
test tube with DNA
polymerase,
nucleotides and short
pieces of synthetic
single-stranded DNA
to act as primers for
DNA synthesis
This is a three-cycle
process which
continues until the
targeted sequence has
been duplicated many
times.
PCR: What is the machine doing?
PCR Process:
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2.
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Heat briefly to separate
DNA strands
Cool to allow primers to
hydrogen bond
DNA polymerase adds
nucleotides to the 3’
end of each primer
After 1 cycle, 2 DNA
molecules are made
Process repeats, each
cycle takes about 5
minutes and doubles
the targeted DNA
sequence each time.
PCR: What is the machine doing?
PCR End Result: millions
of identical DNA
fragments
These copies are used to
produce a DNA profile
which can be used in
crime scene analysis
and paternity testing.
DNA Analysis
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Gel electrophoresis
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Technique used to separate
macromolecules based on
size or electrical charge
Sorts DNA fragments by size
in bands containing
molecules of the same length
Restriction fragment
analysis detects DNA
differences
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Use restriction enzymes to
cut DNA and electrophoresis
to separate fragments
Gel Electrophoresis Process
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2.
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Samples are placed in wells at one end of the gel
Electrodes are attached to electrophoresis chamber
Molecules migrate toward the opposite pole, depending on
charge
When current is turned off, molecules are arrayed in bands
along a lane according to size
Uses of Electrophoresis
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DNA Profiling for
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Paternity Tests
Forensic investigation
DNA Analysis
Human Genome
Project
Time to Practice!
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Electrophoresis Simulation: DNA Goes to
the Races
Human Genome Project
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In 1990, the Human Genome
Project started as an
international collaboration to
determine the entire base
sequence of the human genome.
Public research groups
(universities) and private
companies (Celera) worked on
the project
Completed in 2003
Continue to work on locating
genes and mapping on specific
chromosomes
Human Genome Project
Outcomes:
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Value in knowing the
sequence of genes on
chromosomes, useful in
medicine, forensics and
evolution
Medical benefits:
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Improved diagnosis of disease
Early detection of genetic
susceptibility to disease
Better identification of carriers
of genetic disorders
Drug design to find new
classes of drugs to act on
specific genes
Human Genome Project
Outcomes:
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Improved techniques have
made it possible to find
genomes of other
organisms
Provided new insights into
role of “junk DNA”
(sections of DNA that are
not transcribed)
Found far fewer protein
coding regions than
expected
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Humans have about 25,000
genes
Human Genome Project
Outcomes cont’d
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Knowledge of the significance of
certain sequences
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Presence or absence can be
detected using microarray
technology
New research fields have emerged
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Bioinformatics: use of computers
to store and analyze huge amounts
of data being generated by
sequencing the genome
Pharmacogenomics: links
differences in genomic information
in different populations to
differences in their response to drug
treatment
Cloning Eukaryotic Cells
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A cell is taken from the desired
organism; an egg cell is taken from a
surrogate
The nucleus is removed from the egg
The desired cell and the nucleus free
egg are fused together
The egg grows to become an embryo
Embryo is implanted in the surrogate and
develops into the desired organism
GMOs
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Genetically Modified Organisms: organisms in
which their genetic make up has been altered
Benefits to GM products:
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Crops: enhanced taste and quality; reduced
maturation time; increased nutrients, yields and stress
tolerance; improved resistance to disease, pests and
herbicides
Animals: increased resistance, productivity,
hardiness, and feed efficiency; better yields of meat,
eggs, and milk; improved animal health
Environment: “friendly” bioherbicides and
bioinsecticides; conservation of soil, water, and
energy; better natural waste management; more
efficient processing
Society: increased food security for growing
populations
GMOs
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Controversies:
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Safety: potential human health impact (allergies, transfer
of antibiotic resistance), unknown potential environmental
impact, unknown effect on other organisms, loss of flora
and fauna biodiversity
Access and Intellectual Property: domination of world
food production by few companies; increased dependence
on industrial nations by developing countries;
Ethics: violation of natural organisms’ intrinsic values;
tampering with nature by mixing genes among species;
objections to consuming animal genes in plants and vice
versa; stress for animals
Labeling: not mandated by some countries (US); mixing
GM crops with non-GM crops
Society: new advances may be skewed to interest rich
countries
GMOs
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Current Examples
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*Salt tolerance in tomato
plant
*Synthesis of beta-carotene
in rice (golden rice)
*Factor IX (for human blood
clotting) in sheep milk
Herbicide resistance in crop
plants (round-up ready
plants)
Glowing Bacteria… you will
do this!
(* = Examples from IB, must know 2)
Issues in Biotechnology
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Benefits and harmful
effects of genetic
modification
Ethical Issues of
therapeutic cloning in
humans: creating an
embryo to supply
embryonic stem cells for
medical use
12-13 need more here
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Cloning Eukaryotic Cells