Basic Terms and
Concepts
• Knowledge of DNA has led to an ability to
manipulate the genes of organisms
• We can clone genes and then use them to alter the
genome of viruses and cells
• Known as genetic engineering
• Cloning the
production of
identical copies of an
organism through
asexual means
• Gene cloning the
production of
identical copies of a
single gene
• Accomplished through
• Recombinant DNA
technology
• Polymerase chain
reaction (PCR)
• Reasons for gene cloning
• Produce large quantities of a
gene’s protein product
• Ex: insulin
• Learn how a cloned gene codes for
a particular protein
• Use the genes to alter the
phenotypes of other organisms in a
beneficial way
• When used to modify humans it’s
referred to as gene therapy
• Organisms with foreign DNA or
genes inserted into them are called
transgenic organisms
• Recombinant DNA (rDNA)
• Contains DNA from two or more different sources
• Ex: human cell and bacterial cell
• To make rDNA, a researcher needs a vector
• Vector: a piece of DNA that can be manipulated
such that foreign DNA can be added to it
• Example: plasmid a very small accessory ring of DNA
from bacteria that are not part of the bacterial
chromosome and are capable of self-replicating
• Polymerase Chain Reaction
• Can create billions of copies of a
segment of DNA in a test tube
• Amplifies a targeted DNA sequence
• Requires the use of DNA polymerase
and nucleotides
• Involves 3 basic steps that occur
repeatedly to create many copies of
DNA copies exponentially
• Analysis of PCR
• Each person has a unique collection of
DNA fragment size
• Gel Electrophoresis
• An electrical current is used to force
DNA through a porous gel material
• Fragments are separated according
to their size
• Smaller fragments move farther through
the gel than larger fragments
• Result in a pattern of distinctive bands
called DNA Fingerprints
Types of cellular
cloning
• Besides recombinant DNA, there are two other types of
cloning Reproductive & therapeutic cloning
• Reproductive cloning
• a technology used to generate an animal that has the same
nuclear DNA as another currently or previously existing animal
• Ex: Dolly was a sheep created by reproductive cloning technology.
• Process
• Somatic cell nuclear transfer (SCNT
• Involves the transfer genetic material from the nucleus of a donor adult cell
to an egg whose nucleus, and thus its genetic material, has been removed
• The reconstructed egg containing the DNA from a donor cell must be
treated with chemicals or electric current in order to stimulate cell division.
• Once the cloned embryo reaches a suitable stage, it is transferred to the
uterus of a female host where it continues to develop until birth
• Therapeutic Cloning
• Also called "embryo cloning,”
• The production of human embryos for use in research
• Goal of this process is not to create cloned human beings,
but rather to harvest stem cells that can be used to study
human development and to treat disease.
• Stem cells are important to biomedical researchers because they
can be used to generate virtually any type of specialized cell in
the human body.
• Stem cells are extracted from the egg after it has divided
for 5 days.
• The egg at this stage of development is called a blastocyst
• Medical applications for
Therapeutic cloning
• Treatment for degenerative diseases
•
•
•
•
Alzheimer's’
Stroke and heart diseas
Nerve disorders like Parkinson's,
Disorders involving paralysis or
degeneration of the spine
• Muscular dystrophy
• Demylination ALS, ALD
• Designer cancer therapy
• Generation of “tailor made” organs
• No need for anti-rejection drugs
• Decreases demand for transplant organs
• Application for Organ Transplants
• Known as organogenesis: the growth, or creation, of organs using
stem cells
• To do this, DNA would be extracted from the person in need of a
transplant and inserted into an enucleated egg.
• After the egg containing the patient's DNA starts to divide, embryonic stem
cells that can be transformed into any type of tissue would be harvested
• The stem cells would be used to generate an organ or tissue that is a genetic
match to the recipient
• In theory, the cloned organ could then be transplanted into the
patient without the risk of tissue rejection.
• If organs could be generated from cloned human embryos, the need
for organ donation could be significantly reduced
Transgenic
organisms
• Bacteria, plants, and animals that are
transgenic are more commonly called
genetically modified organisms (GMO)
• Products from GMO’s are called biotechnology
products
• Ex: blood clotting factors, insulin, human growth
hormone, hepatitis B vaccine
• Transgenic Bacteria
• Produced using recombinant DNA
• Grown in vats called bioreactors
• The bacteria express the cloned gene, and the
gene product is usually collected from the
medium where the bacteria is grown
• Uses
• Medical: insulin & vaccines
• Agriculture: use of bacteria to make frostresistant plants, insect-resistant corn
• Bioremediation: naturally occurring bacteria
that eat oil can be genetically engineered to
do an even better job of cleaning up oil spills
• Biochemistry: a certain bacteria is used to
create phenylalanine, an organic chemical
used to make aspartame, an artificial
sweetener
• Transgenic plants
• Foreign genes are placed
into protoplants, or immature
plant embryos
• Will develop into mature plants
that express the foreign DNA
• Example
• Pomato
• Plant produces potatoes below
ground and tomatoes above
ground
• Foreign genes transferred to
cotton, corn ,a dn potato strains
have made them resistant to
pests
• Cells produce insect toxin
• Transgenic animals
• Method of DNA combination: Vortex mixing
• Eggs are placed in an agitator with DNA and slicon-carbide
needles
• The needles make tiny holes in the eggs through with the DNA can enter
• When the eggs are fertilized, the resulting offfspring are
transgenic animals
• Applications
• Eggs have acquired the gene for bovine growth hormone, which
produces larger fish, cows, pigs, rabbits, and sheep
• Gene pharming
• The use of transgenic farm animals to produce pharmaceuticals
• Genes that code for therapeutic and diagnostic proteins are incorporated
into an animal’s DNA, and they appear in the milk
• Examples
• Cystic fibrosis, cancer, and blood diseases
Gene Therapy & the
Human Genome
Project
• Gene therapy is a treatment that involves altering
the genes inside your body's cells to stop disease.
• Genes that don't work properly can cause disease.
• Gene therapy replaces a faulty gene or adds a new
gene in an attempt to cure disease or improve your
body's ability to fight disease.
• Gene therapy holds promise for treating a wide
range of diseases, including cancer, cystic fibrosis,
heart disease, diabetes, hemophilia and AIDS.
• Currently, in the United States, gene therapy is
available only as part of a clinical trial.
• Researchers are testing
several approaches to
gene therapy, including:
• Replacing a mutated gene
that causes disease with a
healthy copy of the gene.
• Inactivating, or “knocking
out,” a mutated gene that is
functioning improperly.
• Introducing a new gene into
the body to help fight a
disease.
• One challenge is gene delivery
how to get the new or replacement
genes into the desired tissues
• Use of a vector is needed
• Most effective vector is the use of
harmless viruses
• Virus DNA is removed and replaced with
the new or replacement genes
• Gene is delivered via the vector to the
host cells
• Stem cells containing the desired gene
can also be used as a vector, which
are incorporated into the affected
tissue
• Example
•Imagine, for example, a little boy
with hemophilia, a condition that is
caused by a faulty gene that makes
his liver unable to make blood
clotting factor 8
• Gene therapy would involve putting a
working copy of the gene which codes
for factor 8 into his liver cells so that
his liver could then produce adequate
levels of factor 8
• Human Genome Project
• an international research effort to sequence and
map all of the genes of members of our
species, Homo sapiens
• known as the genome
• Completed in April 2003
• HGP gave us the ability, for the first time, to read nature's
complete genetic blueprint for building a human being
• Project Goals
• identify all the approximately
20,000-25,000 genes in human
DNA
• determine the sequences of the
3 billion chemical base pairs
that make up human DNA
• store this information in
databases
• improve tools for data analysis
• transfer related technologies to
the private sector
• address the ethical, legal, and
social issues (ELSI) that may
arise from the project.