Download gen-30516-lecture-15-biotech

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Biotechnology is broadly defined as technologies
that involve the use of living organisms, or their
products, to benefit humans
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It is not a new topic
 It began about 12,000 years ago when humans began to
domesticate animal and plants for the production of food
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Since the 1970s, molecular genetics has provided new,
improved ways to make use of organisms to benefit humans
 Genetically modified organisms (GMOs) have received
genetic material via recombinant DNA technology
 An organism that has integrated recombinant DNA into its
genome is called transgenic
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USES OF MICROORGANISMS IN BIOTECHNOLGY
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Microorganisms are used to benefit humans in various ways
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Molecular genetic tools are very important in influencing and
improving our use of microorganisms
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Overall, the use of recombinant microorganisms is an area of
great research interest and potential
 However, there are problems such as safety concerns and
negative public perception
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Many Important Medicines Are Produced by
Recombinant Microorganisms
Insulin
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Insulin regulates several physiological processes,
particularly the uptake of glucose into fat and muscle cells
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Persons with insulin-dependent diabetes have a defect in
their b cells
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Therefore, they cannot synthesize enough insulin
Sources of insulin included
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It is produced by the b cells of the pancreas
Cows
Human cadavers!
But now, patients can use insulin made by recombinant
bacteria – expression vector – eukaryotic gene expressed
in a bacteria
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GENETICALLY MODIFIED ANIMALS
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The production of transgenic animals is a
relatively new, exciting area of biotechnology
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It holds great promise for innovations in
biotechnology
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Of course, this is predicated on public
acceptance!
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Genes can be introduced from different species. Here, different fluorescent
proteins from jellyfish are expressed in zebrafish.
Glo fish
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Transgenic Livestock
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Transgenic species of livestock are being developed
 May include production of medicines in the milk of these
animals
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molecular pharming
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REPRODUCTIVE CLONING
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Reproductive cloning refers to methods that produce
two or more genetically identical individuals
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Cloning is an easier undertaking in plants
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Identical twins are genetic clones from one fertilized egg
Plants can be cloned from somatic cells
For several decades scientists believed that
mammalian somatic cells were unsuitable for cloning
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But in 1997, Ian Wilmut and his colleagues at the Roslin
Institute created Dolly!
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Dolly and her lamb Bonnie
Protocol for the successful
cloning of sheep
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Protocol for the successful
cloning of sheep
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Evidence suggested that Dolly may be “genetically older”
than her actual age would have indicated
 At 3 years old, the length of the telomeres in her somatic
cells were consistent with a sheep that is 9 or 10 years old
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The sheep that donated the somatic cell that produced Dolly
was 6 years old
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6-year old Dolly was euthanized after an examination showed
progressive lung disease
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Her death raised concerns that the techniques used to
produce Dolly could have caused premature aging
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As much as 4% of genes were not expressed normally
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
In recent years, cloning from somatic cells has been achieved
in several mammalian species
 Sheep, cows, mice, goats and pigs
 Unlike the case with Dolly, telomeres in cloned mice and
cattle appear to be the correct length!
 However, other studies have shown other genetic flaws
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With regard to livestock, farmers can use somatic cells from
their best individuals to create genetically homogeneous
herds
 This may be advantageous with regard to agricultural yield
 However, such a herd may be more susceptible to rare
diseases
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Cloned cats and dogs
Cloned cows
Cloned horses
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People have become greatly concerned with the possibility of
human cloning
 To some, it is morally wrong and threatens the basic fabric
of parenthood and family
 To others, it offers a new avenue of reproduction
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For infertile couples who might want a genetically related child
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In the public sector, the sentiment toward human cloning has
been generally negative
 Indeed, many countries have issued an all-out ban
 While others permit limited research in the area
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In the future our society will have to wrestle with the
legal and ethical aspects of cloning
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Stem Cells
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Stem cells supply the cells that construct our bodies
from a fertilized egg
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In the adult, stem cells also replenish damaged cells
Stem cells have two common characteristics
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1. They have the capacity to divide
2. They have the capacity to differentiate into one or more
specialized cell types
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totipotent cells, like fertilized eggs can give rise to all cell types
pluripotent cells can differentiate into almost every cell, but can’t
give rise to an entire, intact individual
multipotent cells can differentiate into several cell types
unipotent cells can only differentiate into one cell type
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When stem cell divides, one may remain undifferentiated,
while the other can differentiate into a specialized cell type
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Thus the population of stem cells remains constant
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In mammals, stem cells are commonly categorized based on
the developmental stage and the ability to differentiate
Can produce all cell types
in an adult organism
Can give rise to
an entire organism
Found in the early
mammalian embryo
Can produce almost all cell
types in an adult organism
However, a single cell
has lost the ability to
produce an entire organism
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Can differentiate into
several cells types
For example,
hematopoietic stem cells
(blood cells) of the bone
marrow
Can only differentiate
into a single cell type
For example, primordial
germ cells in the testis 
sperm, only
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Stem Cells
Adult stem cells are rare
1 cell in 10,000 in the bone marrow is a stem cell
Embryonic Stem cells (ES) and Embryonic Germ cells (EG)
can be grown in the laboratory
 Are easily identified
 Provide greatest potential for transplantation therapy
 Most ES cells are derived from unused embryos from in
vitro fertilization
 Most EG cells are derived from aborted fetuses
 This creates an ethical dilemma in using these cells
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Genetically Modified Plants
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Selective breeding has been used for centuries
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This has produced plants with desirable characteristics
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larger, disease resistance, high-quality food
Genetically engineered crops have been used
since mid-1990s
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In 2009, roughly 25% of all crops were transgenic
More than 100 million hectares planted with transgenics
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Agrobacterium tumefaciens Can Be Used to Make
Transgenic Plants
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The production of transgenic plants is somewhat
easier than transgenic animals
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Certain plant cells are totipotent
 An entire organism can be regenerated from a
somatic cell
Agrobacterium tumefaciens is a bacterium that
naturally infects plants causing crown gall tumor
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Ti plasmid (Tumor-inducing) carried by bacterium
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Transferred DNA
Tumor-inducing
plasmid
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The A. tumefaciens T DNA can be used as a vector
to introduce cloned genes into plants
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First, the Ti plasmid needs to be modified
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The genes that cause tumors are deleted
Selectable marker genes are inserted into the T DNA
R is commonly used
 Kan
Unique restriction sites are added for the convenient
insertion of any gene
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T DNA
integrates into
the genome
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A. tumefaciens does not infect all plant species
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Fortunately, other methods are available
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Biolistic gene transfer (i.e., biological ballistics)
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Microinjection
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The second most common way to produce transgenic plants
A “DNA gun” is used to shoot DNA-coated microprojectiles into the
cells
Microscopic-sized needles are used to inject DNA into the cells
Electroporation
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An electric current is used to create transient pores in the plasma
membrane through which DNA can enter
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Potato plants transformed with a Bt gene
from bacteria
Control
Transformed
(Bt gene)
Genetically engineered plants that are insect
resistant
Herbicide resistant soybeans
Virus resistant transgenic papaya
Golden Rice
vitamin-enriched bananas
The Department of Agriculture has approved
genetically modified apples that never turn
brown
A glowing plant that could provide a sustainable light source
Alfalfa transformants with the maize SPS gene driven by a constitutive promoter
Non transformants
SPS3
SPS6
SPS12 SPS16
SPS1
8
The transformants outperform the non transformed plants significantly
SPS 22
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In the U.S., genetic screening for certain disorders has
become common medical practice
 For example
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Pregnant women over 35 years of age are screened routinely to
see if they are carriers of chromosomal abnormalities
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Rates of such defects increase with the age of the mother
Widespread screening for phenylketonuria
Genetic testing has also been conducted on specific
population in which a genetic disease is prevalent
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Genetic testing can be performed in conjunction
with in vitro fertilization (IVF)
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called Preimplantation genetic diagnosis (PGD)
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IVF involves combining sperm and egg outside of the
mother’s body
One or two cells are removed at 8 cell stage
Tests are done to check for problems
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Molecular tests can check for particular gene defects
Chromosome composition can be checked
Decision can be made whether to implant or not.
Many ethical questions arise from this process
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Pre-implantation genetic diagnosis
Genetic Screening is amongst the newest and most sophisticated
techniques used to test for genetic disorders by direct examination of the
DNA itself.
HUMAN GENE THERAPY
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Gene therapy is the introduction of cloned genes
into living cells in an attempt to cure disease
Research efforts in gene therapy is aimed to
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Alleviate inherited diseases
Treat diseases such as cancer and heart disease
Combat infectious diseases such as AIDS
Human gene therapy is still at an early stage
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Nevertheless, some of the initial results are promising
and future prospects abound
*Gene editing
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Pre-implantation genetic diagnosis
Gene Therapy Involves the Introduction of Cloned
Genes into Human Cells
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Two transfer methods are used
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1. Nonviral approach
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Liposome technique most common
2. Viral approach
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Most common are retroviruses, adenoviruses and parvoviruses
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Aerosol Sprays May Be Used to Treat Cystic Fibrosis
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Cystic fibrosis (CF) is one of the most common
recessive inherited disorders
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Affects about 1 of 3,000 babies of Northern European
descent
It is caused by a defect in a gene that encodes an ion
transport protein
This leads to an abnormality in salt and water balance
This, in turn, leads to accumulation of mucus in the lungs
The result is chronic lung infections which prove fatal
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CF has been the subject of much gene therapy
research
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To implement CF gene therapy, the normal CF gene must
be delivered to lung cells
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The ex vivo approach used in ADA gene therapy is not possible
CF gene therapy has involved the use of an aerosol
spray
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When inhaled by the patient via an aerosol spray, the lung
epithelial cells take up liposomes and adenoviruses
Still at an early stage of development
Eventually may become an effective method of treatment
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Variable-number tandem repeats (VNTRs, or minisatellites)
contain repeating clusters of 10-100 nucleotides that are variable
between individuals
DNA fingerprinting – Forensics
DNA fingerprinting – Paternity tests