Alfred Hershey and Martha Chase (presented by Kristy)

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Transcript Alfred Hershey and Martha Chase (presented by Kristy)

Effects of Bacteriophages on
Modern Biology
By Kristi McKee
Scientific Discovery
Fall 2003
What is a Bacteriophage
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Viruses that attack bacteria
Non-self replicating
Made up of mostly proteins and Deoxyribonucleic
acids (DNA)
Bacteria specific
Inject their genome into host cell
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–
Lytic cycle
Lysogenic cycle
Phage Appearance
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Average length is
around one twenty-fifth
of a micron, but vary in
size.
First use of how phages
can manipulate the
cells production was
producing human
insulin.
What phages do to Host Cell
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www.cellsalive.com
Example of Lytic Life Cycle
http://www.cat.cc.md.us/course
s/bio141/lecguide/unit2/virus
es/rellyso.html
Discovery of Bacteria Infecting Viruses
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Frederick W. Twort
given first credit for
phages.
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Found by studying
micrococcus colonies
Naming of the “Viruses”
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Felix D’ Herelle
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Born in Montreal 1873
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Trained in medical
bacteriology
Came across first in
Mexico with locusts
What D’Herelle Did and Found
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Collected sick locusts and studied the
blackish diarrhea.
In all samples found bacteria locust
coccobacillus
Took healthy colonies and dusted plants that
the insects ate and watched them as well
become infected.
He tested this dust all over the world.
What D’Herelle Did and Found
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Also set up agar plates with colonies of
coccobacilli that was collected from insects.
Made a slide from the agar but saw nothing
under the microscope
Decided that the cause was a filterable. So
he used a porcelain filter that would retain all
bacteria.
The Dawning of the Light
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1915 visiting Paris asked to investigate an
epidemic of dysentery in the Calvary
squadron.
Decided to follow one patient from the time
they were admitted till end of convalescence
to see at what time the principle causing
agent of the clear spots occurred.
Results of Tracking Patient
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1st day normal growth on plate
2nd and 3rd no changes
4th day made emulsion with a few drops from
the bloody stools, filtered through a
Chamberlain candle, added to broth culture
of dysentery bacillus isolated from 1st day.
Filtrated again and spread to a plate and
broth culture. Incubated at 37o.
The Morning After
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The broth culture that was turbid, cleared as
well as the agar spread was also devoid of
all growth.
Went to hospital to share his results and
check on patient.
Reaction of D’Herelle
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"...on opening the incubator I experienced one of those rare
moments of intense emotion which reward the research worker
for all his pains: at first glance I saw that the broth culture,
which the night before had been very turbid was perfectly clear:
all the bacteria had vanished... as for my agar spread it was
devoid of all growth and what caused my emotion was that in a
flash I understood: what causes my spots was in fact an
invisible microbe, a filterable virus, but a virus parasitic on
bacteria. Another thought came to me also, If this is true, the
same thing will have probably occurred in the sick man. In his
intestine, as in my test-tube, the dysentery bacilli will have
dissolved away under the action of their parasite. He should
now be cured"
The Light Comes On
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From all the results D’Herelle now had he
knew it was a filterable virus that was
parasitic on bacteria.
He coined the name bacteriophages
Tried to use phages to treat and prevent
other diseases such as: anthrax, typhus and
bubonic plague.
Short Life of Phage Therapy
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After 1920’s there was a skyrocket in the use
of phage therapy.
Even as late as WWII phage therapy was
being used prevalently in some countries.
However, antibiotics began to phase them
out.
Beginnings of Phage Research
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FM Burnet – origins of phage research
Showed that bacteriophages vary widely in
physical and physiological properties
Different phages carry different antigens
Proposed single-burst method
Discovering Linear Dimensions
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Martin Schlesinger – studied properties of
phages by their chemical and
physiochemistry methods to give first insight
of what a phage may look like.
Showed they are ~0.1mm in length and
~4x10-10g.
Found that phages consisted of mainly
proteins and DNA in equal proportions.
Max Delbruck
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He is often referred to as the founder of
molecular biology.
Perfected a method to culture phages and
found they can infect a cell and with in thirty
minutes lyse cell and have hundredfold their
original number – one step growth method
Along with Hershey found bacteria can
spontaneously mutate to become immune to
phages.
Delbruck continued
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Worked with many other scientists to find that
viruses contain DNA.
As well as helped to reveal the structure of DNA
itself.
Awarded the Nobel for work on phages in 1969
along with Alfred Hershey and Salvador Luria.
Found that phages could also after infecting a cell
phages could exchange genetic information to form
new viral strains
First Electron Micrograph
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Luria and Anderson –
1942 first electron
micrograph picture of a
T2 phage
Anderson also
discovered the phages
adsorbed by the tail by
“critical point” technique
for avoiding surface
tension.
Theoretical goes Experimental
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Through the theoretical contributions of
Delbruck and Luria, Hershey used their
findings to produce his skillful
experimentation along with Martha Chase.
Penetration of Virus to Host Cell
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“Blender Experiment” – 1952 Alfred Hershey
and Martha Chase.
Wanted to demonstrate that DNA not protein
was the genetic material of life.
Early Evidence that DNA is the
Substance of Life
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Miescher – isolated DNA from salmon sperm
found DNA is made up of 4 monomers.
First half of 20th century nucleic acids were
thought to be only structural material in the
cells nucleus.
Hershey and Chase’s experiment was
benchmark in changing ideas.
Focus of Study
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Study involved T2
bacteriophages that
infected Escherichia
Coli.
The “Blender Experiment”
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Wanted to show that the DNA passed on was the
signal for replication and growth
1951 – 52 using radioactive phosphorus and sulfur
they grew phages on media.
Then let phages infect a bacteria culture to replicate.
The process was interrupted by spinning the culture
in an everyday Waring kitchen blender.
The shearing of the blender separated phage protein
from bacteria cells.
“Blender Experiment” cont.
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The cultures were spun in a centrifuge to
separate the liquid from the bacteria that
formed a pellet at the bottom.
Using the tags as a marker for where the
phage protein and DNA was gave the results
and proof they had been hoping for.
DNA, not protein, was the genetic material of
life.
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www.accessexcellemce.org
Leads for Future Experiments
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Scientist now wanted to know how DNA:
Operated
Replicated
Direct the production of proteins
What was its structure
Results from Learning of Phages
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1970 Hamilton Smith found first site-specific
restriction enzyme using phages.
Antibiotic resistant bacteria: use of phages to destroy
bacteria.
Search for new potential drugs.
Develop human antibodies.
Biosensors
Only ~0.0002% of global phage genome has been
sampled.
Resources
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“Alfred Day Hershey”, Discovering Science Bibliography
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“Max Delbruck”, Discovering Science Bibliography
“T4 Bacteriophages attacking an E. Coli Bacterium”,
Discovering Science
Evers, Chris. “Experiments that Inspire”
www.acessexcellence.org
Mathews, Christopher K.; VanHolde, KE.; Biochemistry
2nd Ed. Benjamin/Cummings Publishing Company, Inc,
Menlo Park Ca. 1996. pp 93, 94.
“Max Delbruck – Nobel Lecture” www.nobel.se
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Recourses Cont.
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“The Nobel Prize in Physiology or Medicine 1969”
www.nobel.se
Rohwer, F. 2003 “Global phage diversity”. Cell 113 (April 18)
:141.
Seaman, Tracy and Trollip, Andre and Mole, Richard and
Albert, Heidi (2003) “The use of a novel phage-based
technology as a practical tool for the diagnosis of tuberculosis
in Africa”. African Journal of Biotechnology 2(2):40-45.
Travis, John. “All the World’s a Phage” Science News (2003)
Vol. 164(2)