Download Unit 6 Part 2 Notes Jan 16 2012

Advances in Genetic Technology
Class Notes
Make sure you study this along
with our first PowerPoint on
Transgenics and your class Article
notes
When we talk about cloning, are
we always talking about the same
thing?
• recombinant DNA cloning (transfer of gene of
interest to a vector like bacteria or a virus
which will incorporate in all future DNA)
• reproductive cloning (transfer of nuclear
material from a somatic cell of one organism to
an egg of another that has had its nucleus
removed, resulting in an offspring like the
donor)
• therapeutic cloning (also known as embryo
cloning is designed to create stem cells for use
in treating disease)
How is a mammal
cloned (reproductive
cloning)?
Is this possible In
humans?
There are lots of
efficiency problems.
It takes hundreds of
tries to get one
clone, even in a
sheep.
What is PCR and why do we need
it?
• Polymerase chain reaction (PCR) is a fast and
inexpensive technique used to "amplify" - copy small segments of DNA.
• It is used wherever large amounts of DNA are
needed. Most often there is only a small sample
of DNA to begin with.
• Examples of its use: DNA fingerprinting, detection
of bacteria or viruses (particularly AIDS), and
diagnosis of genetic disorders.
Steps to PCR
mimic what
happens in
replication
If you run about 35 cycles and end up with
billions of copies of the original DNA
What is a Short Tandem Repeat
(STR)Polymorphism and how is it used?
• STRs are short sequences of DNA, normally of length
2-5 base pairs, that are repeated numerous times.
• These are inherited regions of our DNA that can vary
from person to person.
• These are located at the same place on the
chromosomes but they vary (polymorphism) in
length from person to person.
• It is highly unlikely that an individual would have the
same number of repeats as another individual at the
most common13 STR sites used by criminal justice to
distinguish individuals.
These loci on the chromosomes are the location of
STRs and are used to distinguish one individual from
another. PCR can be used to make many copies of the
DNA segments with STRs for forensic analysis.
Electrophoresis is used in separating DNA
fragments
• Gel electrophoresis is a method of separating large molecules
(such as DNA fragments or proteins) from a mixture of similar
molecules.
• An electric current is passed through a gel containing the DNA
mixture, and each kind of molecule travels through the medium
at a different rate, depending on its electrical charge and size.
• The gel acts as a sieve. DNA moves at different rates because
larger molecules move through the gel matrix slower than smaller
molecules.
The Human Genome Project
(approx. 1990-2003)
Project goals were to:
• 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
Applications of the Data from the
Human Genome Project
• Molecular medicine
• Energy sources and environmental
applications
• Risk assessment
• Bioarchaeology, anthropology, evolution, and
human migration
• DNA forensics (identification)
• Agriculture, livestock breeding, and
bioprocessing
What is Gene Therapy?
Gene therapy is designed to introduce beneficial versions of a
gene into an afflicted individual for therapeutic purposes.
How does it work?
• For gene therapy to be permanent, the cells that receive
the normal allele must be ones that multiply throughout
the patient's life.
• In most gene therapy studies, a "normal" gene is inserted
into the genome to replace an "abnormal," disease-causing
gene using a vector.
• Currently, the most common vector is a virus that has been
genetically altered to carry normal human DNA.
Two mechanisms: direct into the tissue or through the use
of stem cells (unspecialized cells) to carry the gene into the
body.
How do small RNAs (microRNA, RNAi)
impact the expression of traits?
• These are short nucleotide sequences that
bind to the complementary RNA made during
transcription and they usually “silence” the
gene.
• MicroRNAs target about 60% of all genes.
They are abundant in all human cells and are
able to repress hundreds of targets each.
When the microRNA interferes with translation the protein
will not be produced and therefore the gene is “silenced” or
not expressed.
How can we determine if a gene will “express”
itself?
With microarray technology.
A microarray is a tool for analyzing gene expression that
consists of a small membrane or glass slide containing samples
of many genes arranged in a regular pattern.
Scientists can determine, in a single experiment, the
expression levels of hundreds or thousands of genes within a
cell by measuring the amount of mRNA bound to each site on
the array.
Often they are looking for Single Nucleotide Polymorphisms
(SNPs)
Compare a normal healthy sample with the
patients sample
GREEN represents Control DNA,
RED represents Sample DNA, where either
DNA or cDNA is derived from diseased tissue
hybridized to the target DNA.
YELLOW represents a combination of
Control and Sample DNA, where both
hybridized equally to the target DNA.
BLACK represents areas where neither the
Control nor Sample DNA hybridized to the
target DNA.
Different types of microarrays have different
applications
In Brief: Microarray Applications
Microarray type
Application
CGH
Tumor classification, risk assessment,
and prognosis prediction
Expression analysis
Drug development, drug response,
and therapy development
Mutation/Polymorphism analysis
Drug development, therapy
development, and tracking disease
progression
What is a SNP?
• It is a specific type of mutations in DNA. This small variation is
enough to cause a disease or disability.
• When researchers use microarrays to detect mutations or
polymorphisms in a gene sequence, the target, or
immobilized DNA, is usually that of a single gene.
• In this case though, the target sequence placed on any given
spot within the array will differ from that of other spots in the
same microarray, sometimes by only one or a few specific
nucleotides.
• One type of sequence commonly used in this type of analysis
is called a Single Nucleotide Polymorphism, or SNP, a small
genetic change or variation that can occur within a person's
DNA sequence.
Last but not least, what does epigenetics have to
do with this?
• Epigenetics influence the expression of traits.
• It is now clear that your destiny is not limited by your
genes alone. There are plenty of non-genetic influences
on expression of your traits.
• It is through epigenetic marks that environmental factors
like diet, stress and prenatal nutrition can make an
imprint on genes that are passed from one generation to
the next
•Epigenetic mechanisms include methylation and
acetylation of nucleotides which cause the DNA to bind
tightly or more loosely around the histones.
•The ultimate result is the transcription of information
onto mRNA or not.