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Gene Technologies
Analyzing Genes
• Multiple technologies exist to analyze
genes of individuals or even populations,
including:
• Gel electrophoresis
• DNA microarrays
Gel Electrophoresis
• Electrophoresis can be used to analyze
DNA, RNA, or proteins.
• The polarity of these molecules causes
them to move a particular direction
through a gel when an electrical current
is passed through it.
• The size of the DNA, RNA, or protein
fragments affects the rate of movement.
The properties of DNA allow it to be extracted easily from
living tissue. DNA is soluble in water, so cells are ground
up in water first. DNA is insoluble in alcohol, so a layer of
alcohol can be floated on top of the solution and DNA
precipitates out of solution at the water-alcohol interface.
The crude DNA extraction also contains fragments of proteins
and other molecules, so it has to be purified. Once purified, it
can be cut with enzymes to remove just the segments of
interest. Then the DNA fragments are put in a PCR machine
with free nucleotides and DNA polymerase. Cycles of heat (to
unzip DNA but not denature the enzyme) and cool (to allow
DNA to “zip” together) cause multiple copies of the DNA to be
made.
The purified and amplified DNA fragments are loaded
into a gel, usually made either of agar (a carbohydrate)
or polyacrylamide (a synthetic molecule). The gel is
covered with a buffer solution and an electrical current is
run through the gel. The DNA fragments migrate down
the gel, which is later stained or put under UV light to
show DNA bands.
DNA Microarrays
• Electrophoresis can usually only test a
few genes or individuals at a time.
• New DNA microarrays (“gene chips”) can
test for thousands of genes at once.
Microarrays take advantage of the base-pairing rule.
Small DNA fragments can be manufactured to match
genes of interest and printed directly on a silicon chip.
DNA or RNA may extracted from the organism of interest
– DNA to analyze all genes present, RNA if the
researcher is interested in what genes are being
expressed. The extracted material is tagged with
fluorescing dyes, loaded on the chip, and allowed to
hybridized with the probes. The chip is scanned to see
which spots on the array glow, and in what color.
The results may tell a researcher which genes are being
expressed in different cells. For example, cancerous cells
may be compared with noncancerous cells to indentify
which genes are “on” and which are “off,” in search of
which genes cause a cell to turn cancerous.
Changing Genes?
• Some gene technologies involve
changing the genes of an individual.
• We’re going to look at some of the
science and consider some of the risks
and benefits of these technologies.
Harnessing Nature
• Gene-altering technologies often
harness the ability of certain microbes to
insert or alter genes of other organisms.
Bacteria already do “gene therapy” on one another.
(b) Transformation
with DNA fragment
bacterial
chromosome
(c) Transformation
with plasmid
bacterial
chromosome
DNA
fragments
DNA fragment is
incorporated into
chromosome.
plasmid
Plasmid replicates
in cytoplasm.
Viruses are expert at inserting their genes into
living hosts.
virus
viral DNA
2 Virus enters host cell.
host cell
3 Virus releases its DNA into host cell;
some viral DNA (red) may be incorporated
host cell
1 Virus attaches to
DNA
into the host cell’s DNA (blue).
susceptible host cell.
viral DNA
Viral proteins
“hybrid virus”
6 Host cell bursts,
releasing newly assembled viruses.
When “hybrid viruses” infect a
second cell, they may transfer
5 New viruses assemble;
genes from the first cell to the
host cell DNA is carried
second cell.
by “hybrid viruses.”
4 Viral genes encode synthesis
of viral proteins and viral gene
replication. Some host cell DNA
may attach to replicated viral
DNA (red/blue).
“Gene Gun”
• Another method of inserting genes into
cells is the biolistic particle deliver
system, or “gene gun.”
• The “gene gun” uses compressed air to
“shoot” microscopic particles coated with
DNA into cells.
“Gene gun” technology
Gene Therapy
• Gene therapy may provide ways to treat
single-gene genetic disorders.
• Gene therapy takes advantage of viruses
as vectors for inserting “good” genes into
cells that have “broken” genes.
• Ashanti deSilva was
one of the first people
to undergo gene
therapy.
• Ashi was born with
ADA deficiency. She
is missing an enzyme
critical for her immune
system.
• The enzyme was necessary for white blood cell function.
Researchers used a virus that parasitizes white blood
cells to insert a functional gene for the ADA enzyme
• The virus inserted the gene in Ashi’s white blood cells,
where it successfully produced the ADA enzyme
• White blood cells live
a few months, so Ashi
has to return for
frequent treatments. If
her bone marrow cells
could be treated, she
might be cured.
• With gene therapy,
Ashi lives a healthy,
productive life.
• However, there are problems and ethics
involved in gene therapy.
• The therapy is only useful for single-gene
disorders where the affected tissue is
local and easily reached.
• In one study on treating cystic fibrosis
with gene therapy, a good gene was
inserted into a cold virus. A volunteer who
was dosed with the cold virus had a
violent immune system reaction and died.
The study was halted until a safe dose
could be established.
GMOs
• For thousands of years, farmers have been
changing the genetics of their crops through
selective breeding. In the 18th century, after
the discovery of pollination, hybridization was
added as a means of improving crops.
• Only recently have humans been able to
change the genetics of crops and livestock by
deliberately inserting new genes to make
Genetically Modified Organisms (GMOs).
DNA including Bt gene
Ti Plasmid
Cut both with the same restriction enzyme.
Genes are prepared for insertion into a DNA plasmid from
bacteria, which will be used to insert the gene into a plant
cell.
Mix Bt gene and plasmid; add DNA ligase to seal DNA.
The enzyme ligase is used to seal the trans gene into the
bacterial plasmid.
Transform Agrobacterium tumefaciens with recombinant
plasmid.
A. tumefaciens
bacterial
chromosome
plasmids
Plasmids are applied to a culture of bacteria that are known
to infect plant cells.
Infect plant cell with transgenic bacterium.
plant
chromosomes
A. tumefaciens
plant cell
The bacteria attacks a plant cell and attempts to insert its
own DNA. It inserts the trans plasmid instead.
plant
chromosomes
A. tumefaciens
plant cell
Insert Bt gene into plant chromosome.
Bt gene
If all goes well, the gene will be inserted into the plant’s
DNA and will be expressed in the plant.
Transgenic plant cells are treated with hormones to grow
new plants, and plants are tested for expression of the new
gene.
• Successful crops that have been developed by
genetic modification:
• Herbicide-resistant crops
• Crops that produce substances toxic to
insect pests but not to people
• Bananas that contain vaccine proteins, to
vaccinate children in developing nations
against cholera
• Goats have been developed that produce
pharmaceutical proteins in their milk.
GMOs and ethics
• Producing GMO crops is expensive, and
companies expect return on their
investment. Companies patent the
genome of their crops to protect their
investment.
• Consider the following “worst case”
scenarios (based on actual events) and
the ethical issues involved.
• A farmer in Canada grows non-GMO corn.
He saves seed from each crop and plants it
the next year. Technicians from a large
corporation remove corn from his land
without his knowledge, test it, and find that
his corn has cross-pollinated with their
patented GMO corn, which another farmer
several miles away is growing. The
corporation sues the farmer for pirating their
crop. The farmer also loses his organic
status for producing corn that is genetically
modified.
• A large corporation develops a GMO rice
that has more protein and vitamins than
ordinary rice. They give some to rice
farmers in India. These farmers normally
save seeds each year and replant them.
After their first harvest, the corporation tells
the farmers that they must buy seeds next
year, because harvested seeds will be
sterile. The farmers protest. They say they
can’t make a living if they have to buy
seeds each year, and they should have
been told this to begin with.