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Chapter 12: DNA Technology and the Human Genome

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Title: Chapter 12: DNA Technology and the Human Genome


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Chapter 12DNA Technology and the Human Genome
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Scientists show precise control in genetic
engineering of sheep July 3, 2000 By Rick
Weiss / Washington Post Achieving a coveted
scientific first, researchers in Scotland have
developed amethod for cloning large animals that
are endowed precisely with valuable genes or lack
specific undesirable genes manipulations that
until now have been possible only in mice. The
advance makes possible the creation of sheep,
cows or other large mammals that are born with
human genetic diseases for study, and is expected
to speed the development of cloned pigs bearing
"humanized" organs for transplantation into
people. And in theory, at least, the technique
could provide a convenient way to engineer
special traits into human cells, or even into
human clones. "We are clearly at the dawn of a
new era in mammalian genetic technology, Milind
Suraokar and Allan Bradley write in a commentary
accompanying the research report in the current
issue of the journal Nature. More than 10 years
have passed since scientists first learned how to
make specific genetic alterations in mice, in
some cases shooting new genes into
mouse chromosomes and other times "knocking out"
specific mouse genes to see what happens. The
method has revolutionized the study of human
genetic diseases and has made the mouse the
workhorse of the biotechnology era. But for many
purposes the mouse is a less than ideal animal
"model" for human disease, because either it's
too small to study well or the relevant
physiology is so different than in people. For
example, scientists for years have been creating
mice harboring the human gene defect that causes
cystic fibrosis, a disease that mostly affects
the lungs of its human victims. But mouse lungs
are far smaller and quite different from human
lungs. The sheep respiratory system is very
similar to the human equivalent so similar
that some researchers have studied the effects of
cigarettes by forcing sheep to smoke through
masks. The new method could quickly lead to the
development of sheep endowed with the human
cystic fibrosis gene defect, allowing
unprecedented studies of that disease's molecular
underpinnings and accelerating tests of
experimental drugs, said Alan Colman, research
director for PPL Therapeutics, the
Edinburgh-area biotechnology company that
conducted the new work. The process isn't
efficient. Of more than 400 embryos made, only 14
lambs were born. Only six of those were alive a
week after birth, and three of those died within
six months, mostly from birth defects apparently
caused by the cloning process. Nevertheless,
tests showed that all but one of the 14 liveborn
lambs containedthe desired gene in exactly the
desired chromosomal slot. PPL is interested in
creating pigs whose organs look less piglike and
more humanlike, in hopes that the organs will be
acceptable to the human immune system. Already
the team has knocked out one of three key genes
in pig cells known to cause organ rejection. They
have yet to add any of the human genes that
may also be needed, or to clone a pig from such a
gene-altered cell.
3
Biotechnology is a useful tool that is delivering
improved products and will make improved
pharmaceutical, agricultural and industrial
products available. Biotechnology promises a
range of benefits for people around the
world. For example, the following are some of
the benefits of biotechnology in the opinion of
leading scientists and other experts The
Environment. Discoveries in biotechnology allow
for certain key crops to have their own
protection against insects and disease. These
crops can, therefore, be grown using less crop
protection chemicals. This allows farmers to
choose the best combination of products and
farming practices to control harmful pests and
diseases. Feeding a Growing Population. World
hunger is a complex issue and biotechnology is
among the approaches to help address this
challenge. Thanks to developments in food
biotechnology, we'll be able to grow more food
and better food on land already being farmed.
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Nutrition, Quality and Taste of Food. The list
of foods available which are enhanced by
biotechnology is growing. Today, research into
numerous improved food products includes food
crops with higher levels of nutrients that may
help reduce the risk of heart disease and certain
cancers. Medicines and Health Care. Besides
foods, biotechnology is bringing breakthroughs in
health care. Since the initial production of
human insulin to better treat diabetes,
biotechnology continues to create more effective
drugs and vaccines. These medicines benefit
hundreds of millions of people worldwide who
suffer from devastating diseases such as heart
disease, cancer, diabetes, Parkinson's,
Alzheimer's and AIDS. Industrial Applications.
In the future, some applications of
biotechnology will be used to make materials such
as fibers for clothes from "renewable" resources
like corn. Other applications may help reduce our
dependence on oil and natural gas and could
reduce water and energy use by as much as 50
percent.
5
Recombinant DNA technology refers to the set of
techniques for recombining genes from different
sources in vitro and transferring this
recombinant DNA into a cell where it may be
expressed. These techniques were first developed
around 1975 for basic research in bacterial
molecular biology, but this technology has also
led to many important discoveries in basic
eukaryotic molecular biology. Such discoveries
resulted in the appearance of the biotechnology
industry. Biotechnology refers to the use of
living organisms or their components to do
practical tasks such as The use of
microorganisms to make wine and cheese.
Selective breeding of livestock and crops.
Production of antibiotics from microorganisms.
Production of monoclonal antibodies.
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Using recombinant DNA techniques, modern
biotechnology is a more precise and systematic
process than earlier research methods. It is also
a powerful tool since it allows genes to be moved
across the species barrier. Using these
techniques, scientists have advanced our
understanding of eukaryotic molecular
biology. The human genome project is an
important application of this technology. This
project's goal is to transcribe and translate the
entire human genome in order to better understand
the human organism. A variety of applications
are possible for this technology, and the
practical goal is the improvement of human health
and food production.
7
An overview of how bacterial plasmids are used to
clone genes for biotechnology.
8
Restriction Enzymes Restriction enzymes are
major tools in recombinant DNA technology. First
discovered in the late 1960's, these enzymes
occur naturally in bacteria where they protect
the bacterium against intruding DNA from other
organisms. This protection involves restriction,
a process in which the foreign DNA is cut into
small segments. Most restriction enzymes only
recognize short, specific nucleotide sequences
called recognition sequences. They only cut at
specific points within those sequences.
9
Using a restriction enzyme and DNA ligase to make
recombinant DNA. The restriction enzyme in this
example (called EcoRI) recognizes a specific
six-base-pair sequence and makes staggered cuts
in the sugar-phosphate backbone within this
sequence.
10
Plasmids are small loops of DNA found in bacteria
that are used as vectors to carry genes in
recombinant DNA research.
11
Using the Ti plasmid as a vector for genetic
engineering in plants.
3. As the plant cell divides, each of its
descendants receives a copy of the T DNA and any
foreign genes it carries. If an entire plant is
regenerated, all its cells will carry--and may
express--the new genes.
1. The Ti plasmid is isolated from the bacterium
Agrobacterium tumefaciens, and a fragment of
foreign DNA is inserted into its T region by
standard recombinant DNA techniques.
2. When the recombinant plasmid is introduced
into cultured plant cells, the T DNA integrates
into the plant's chromosomal DNA.
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WASHINGTON (Reuters) -- Green glowing mice that
carry a jellyfish gene have shown that sperm can
be used to genetically engineer animals,
researchers said Thursday. The research team --
which made headlines last year by cloning
generations of mice -- said its technique
improved on current hit-and-miss methods in which
scientists painstakingly inject new genes into
animals they are trying to alter or use a virus
to deliver the new genes. Anthony Perry and
colleagues at the University of Hawaii in
Honolulu said they recruited nature's
tried-and-true method for mixing up genes,
using sperm. "The great thing about this new
method is that it is very straightforward," Perry
told Science magazine, which published the
findings. "It may prove to be more
efficient." The same team, which includes
Japanese researchers Teruhiko Wakayama and Ryuzo
Yanagimachi, not only cloned the first mice, but
has done experiments with mouse sperm, using
freeze-dried sperm to fertilize eggs. They
showed that "dead" sperm heads still carried
viable genetic material. In the latest
experiment they took this research a step
further, freeze-drying mouse sperm, mixing them
with DNA from another species, and injecting the
mixture into mouse eggs. They then put those eggs
into female mice using standard techniques for
making test-tube babies in animals
and humans. In this case they used genes that
make jellyfish glow green. It made it easy to
look for the genes in the resulting baby mice --
they just put them under an ultraviolet light to
see if their skin glowed green. "It was
absolutely fantastic to see these little mice
glowing green," Perry said in a telephone
interview. The new pups are bald so the green
glow is easy to detect.
Now. Glowing Mice!
13
News item-
Scientists in Japan have implanted into mice the
gene that makes jellyfish glow. They hope to use
the technique to track various kinds of cellular
activity in organisms. The transgenic mice
radiate an eerie green light. It's very
beautiful,' says chief researcher Masaru Okabe.
What can be done with mice can presumably be done
for humans, too. Will glowing humans become all
the rage, the latest twist in our maddened search
for the ultimate cosmetic, this one built into
the very fabric of the genes? In the meantime,
one can imagine those lovely luminescent mice,
scampering about the house at night, betrayed to
the cat by their jellyfish glow, or going out
like a sputtering candle beneath the sprung wire
of the trap.
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DNA technology is reshaping the medical and
pharmaceutical industries Diagnosis of
Diseases Medical scientists currently use DNA
technology, in particular PCR and labeled DNA
probes, to detect genetic diseases and identify
heterozygote carriers. This allows early disease
detection and identification of carriers for
potentially harmful recessive mutations even
before the onset of symptoms. Genes have been
cloned for many genetic disorders including
hemophilia, phenylketonuria, cystic fibrosis, and
Duchenne's muscular dystrophy. Gene cloning
permits direct detection of gene mutations. A
cloned normal gene can be used as a probe to find
the corresponding gene in cells being tested the
alleles are compared with normal and mutant
standards usually by RFLP analysis.
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Human Gene Therapy Traceable genetic disorders
in individuals may eventually be correctable.
Theoretically, it should be possible to replace
or supplement defective genes with functional
normal genes using recombinant DNA techniques.
Correcting somatic cells of individuals with
well-defined, life-threatening genetic defects
will be the starting place.
19
  • The principle behind human gene therapy is that
    normal genes are introduced into a patient's own
    somatic cells. For this therapy to be effective,
    there are several requirements
  • The few cells receiving the normal gene must
    actively reproduce, so the normal gene will be
    replicated.
  • The cells' normal protein products can correct
    the disorder.
  • Gene therapy is especially suited for
    single-enzyme deficiency diseases.
  • For example, several children with an
    immunodeficiency disease are currently being
    treated successfully by gene therapy their
    disorder results from a lack of the enzyme
    adenosine deaminase (ADA).

20
One type of gene therapy. In this procedure, a
disarmed retrovirus is used as a vector to
introduce a normal allele of a gene into the
cells of a patient who lacks it.
21
Despite its promise, however, gene therapy raises
difficult social and ethical questions Who
will have access to gene therapy? The procedures
are very expensive and only available in major
medical centers. Should we try to treat germ
cells with the hope of correcting a defect in
future generations? Some critics feel that
tampering with human genes is wrong regardless of
the situation, because it will lead to eugenics.
Others say that genetic engineering of somatic
cells is no different than other conventional
medical interventions used to save lives.
22
  • Forensic Uses of DNA Technology
  • Forensic labs can determine blood or tissue type
    from blood, small fragments of other tissue or
    semen left at the scene of violent crime. These
    tests, however, have limitations
  • They require fresh tissue in sufficient amounts
    for testing.
  • This approach can exclude a suspect but is not
    evidence of guilt, because many people have the
    same blood type or tissue type.
  • Theoretically, DNA testing can identify an
    individual with certainty, since everyone's DNA
    base sequence is unique (except for identical
    twins). The most commonly used DNA technology
    for this forensic application is the RFLP
    autoradiograph.

23
This method, known as RFLP analysis, is used to
compare DNA samples from the suspect, the victim,
and a small amount of semen, blood or other
tissue found at the scene of the crime.
Restriction fragments from the DNA samples are
separated by electrophoresis radioactive probes
mark the bands containing RFLP markers. Even a
small set of RFLP markers from an individual can
provide a DNA fingerprint that is of forensic
use the probability that two individuals would
have the same RFLP markers is quite low.
24
A DNA Fingerprint can be used to identify the
perpetrator of a crime through RFLP analysis.
25
DNA technology raises important safety and
ethical questions When scientists realized the
potential power of DNA technology, they also
became concerned that recombinant microorganisms
could create hazardous new pathogens, which might
escape from the laboratory. In response to these
concerns, scientists developed and agreed upon a
self-monitoring approach, which was soon
formalized into federal regulatory
programs. Today, governments and regulatory
agencies worldwide are monitoring the
biotechnology industry - promoting potential
industrial, medical, and agricultural benefits,
while ensuring that new products are safe. In
the U.S., the FDA, National Institutes of Health
(NIH) Recombinant DNA Advisory Committee,
Department of Agriculture (USDA), and
Environmental Protection Agency (EPA) set
policies and regulate new developments in genetic
engineering.
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While genetic engineering holds enormous
potential for improving human health and
increasing agricultural productivity, new
developments in DNA technology also raise ethical
concerns. For example, mapping the human genome
will contribute to significant advances in gene
therapy, but Who should have the right to
examine someone else's genes? Should a person's
genome be a factor in their suitability for a
job? Should insurance companies have the right
to examine an applicant's genes? How do we
weigh the benefits of gene therapy against
assurances that the gene vectors are safe?
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For environmental problems such as oil spills,
genetically engineered organisms may be part of
the solution, but what is their potential impact
on native species? For new medical products,
what is the potential for harmful side effects,
both short-term and long-term? New medical
products must pass exhaustive tests before the
FDA approves it for general marketing.
Currently awaiting federal approval are
hundreds of new genetically engineered diagnostic
products, vaccines, and drugs - including some to
treat AIDS and cancer.
28
For agricultural products, what are the potential
dangers of introducing new genetically engineered
organisms into the environment? Some argue that
producing transgenic organisms is only an
extension of traditional hybridization and should
not be treated differently from the production of
other hybrid crops or animals. The FDA holds
that if products of genetic engineering are not
significantly different from products already on
the market, testing is not required. Some are
concerned that genetically altered food products
may contain new proteins that are toxic or cause
severe allergies genetically engineered crop
plants could become superweeds resistant to
herbicides, disease and insect pests and
engineered crop plants may hybridize with native
plants and pass their new genes to closely
related plants in the wild.
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