Download Biotechnology

Biotechnology
Assessment statements
4.4.1 – 4.4.13
4.4.1 Polymerase Chain Reaction
• A technique that amplifies (clones) a small
amount of DNA into millions of copies.
• Requires a very small initial sample and a
few hours time
The Materials
• Target DNA (your sample you want to
amplify)
• Taq polymerase (DNA polymerase isolated
from a bacteria, Thermus aquaticus in a
thermal spring in Yellowstone)
• DNA primers
• DNA nucleotides
• Taq polymerase remains active despite
repeated heating and cooling.
• The ingredients are added to a test tube
and placed into a thermal cycler that
uses a computer to control the repetitive
temperature changes required for PCR
The Methods
1. Melting – the mixture is heated to 94-96 oC for
a minute or so to denature DNA into single
strands
2. Annealing – the mixture is cooled to 50-65 oC
and primers attach (by H bonds) to their
complementary sequences on either side of
the target sequence
3. Elongation – the mixture is brought to 72 oC for
several minutes during which polymerase
binds and extends a DNA complement from
each primer
A video
• http://faculty.plattsburgh.edu/donald.slish/
PCRmov.html
• After amplification the products of PCR are
loaded into wells of an agarose gel and
electrophoresed.
Gel Electrophoresis
• DNA is chopped into fragments using
restriction enzymes (more on this later)
• Fragments are loaded in a gel that has
small wells at one end.
• The gel is placed in a box that can be
connected to an electric current
• The wells with the DNA are loaded at the
negative end of the box and the current is
turned on
• Because the phosphate groups on DNA
are negatively charged the DNA migrates
through the gel toward the positive pole
• The gel acts as a molecular sieve,
separating the pieces of DNA by size.
An animation
• http://www.sumanasinc.com/webcontent/a
nimations/content/gelelectrophoresis.html
A simulation
• http://learn.genetics.utah.edu/content/labs/
gel/
Sample Results
Restriction Enzymes
• Restriction enzymes (endonucleases) cut
DNA at locations called palindromic
sequences
• These cuts may result in either blunt or
sticky ends
• Different DNA cut with the same restriction
enzyme can be spliced together because
they have the same sticky ends! This
makes genetic engineering possible.
DNA Cloning
• Cloning employs plasmids, small circles of DNA
found in prokaryotic cells that are supplemental
to the bacterial cells main DNA
• Plasmids are removed from host cells and cut
with restriction enzymes. The gene to be copied
is mixed with the cut plasmids and
complimentary ends align. DNA ligase joins
them together. The plasmid is now referred to as
a recombinant and can be used as a vector.
• The vector is inserted in a host bacterium
and it is placed in a bioreactor.
• The host reproduces and copies the gene
• The host also expresses the gene and
synthesizes the protein that the gene
codes for.
• Bacteria can be created to make human
proteins. Ex.) insulin
DNA Profiling
• Matching an unknown sample of DNA to a
known sample. AKA DNA fingerprinting
• Utilizes Satellite (VNTR-variable number
of tandem repeats) DNA - highly repetitive
sequences of DNA from the non coding
region of the genome.
• Different individuals have a unique length
to their satellite regions that can be used
to differentiate between individuals.
An article on DNA profiling
• http://learn.genetics.utah.edu/content/labs/
gel/forensics/
A Simulation
http://www.biotechnologyonline.gov.au/popu
ps/int_dnaprofiling.html
Uses
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Crimes
Paternity
Selective Breeding – endangered species
Animal migration
Establishing evolutionary relationships
Click 4 biology
• http://click4biology.info/c4b/4/gene4.4.htm
#four
• Check out this link for application of this
technology and to see real gels!
Human Genome Project
• After watching “Cracking the Code of Life”,
what ethical issues regarding the human
genome concern you?
• http://www.pbs.org/wgbh/nova/genome/
Exploration
• At the website complete the following
activities:
– Take the survey. Submit your answer to see
how others voted on each issue.
– Manipulating genes (read both parts 1 and 2)
– Understanding heredity (make a list of the
people and each ones major contribution)
– Explore a stretch of code (define hitchhiking
code, ancient code, sites of variation)
– Sequence for yourself (complete the activity)
On your own time
• Nature vs. Nurture
• Journey into DNA
• Meet the decoders
Genetically Modified Organisms
• GMO’s have had an artificial genetic
change using genetic engineering
techniques such as gene transfer or
recombinant DNA
Transgenic Plants
• An undesirable gene is removed and sometimes
a desirable gene is put in its place.
• Scientists can create “designer plants” that
exhibit greater productivity, resistance to pests,
higher nutrient content etc…
• See Click 4 biology for examples and links!
• http://click4biology.info/c4b/4/gene4.4.htm#nine
Transgenic Animals
• New genes can be inserted into animals
so that they produce products that humans
need.
• Ex.) sheep/cows may produce milk that
contains clotting factor for those with
hemophilia.
Pros and Cons of Genetically
Modified Organisms
• Pros – more nutritious, productive, resistant
crops; ability to produce rare proteins for
medicines or vaccines; faster than selective
breeding at giving desired traits
• Cons – genetically modified crops may pollinate
wild species; crops may harm humans through
toxicity or by causing allergy; may lead to a
decline in biodiversity as engineered organisms
increase in number.
Info on GMO’s
• http://www.csa.com/discoveryguides/gmfood/ove
rview.php Mainly deals with crops.
• http://www.actionbioscience.org/biotech/sakko.ht
ml More pros and cons
• http://www.who.int/foodsafety/publications/biotec
h/20questions/en/ World Health Organization
(available in several languages!)
Cloning
• A Clone is a group of genetically identical
organisms or group of cells artificially
derived from a single parent.
Dolly!
Why she was unique!
• Dolly was the first clone whose genetic
information did not originate from an egg!
– An udder cell was collected and cultured. A cultured
cell was selected and its nucleus was removed.
– An unfertilized egg was collected and its nucleus was
removed.
– The egg and the somatic nucleus were fused.
– The new cell began embryonic development and was
implanted in a surrogate mother
– Dolly was born! A clone of the udder cell donor!
Therapeutic Cloning
• Uses undifferentiated cells, often derived
from embryos
• http://www.dnalc.org/resources/animations
/stemcells.html
Stem Cell Webquest
• http://outreach.mcb.harvard.edu/teachers/
Summer07/JimDixon/SC_Biology_webque
st.doc
Homework
• Page 110, #14-17