History_of_DNA.pptDNA History presentation ppt

DNA History presentation ppt

2. Griffith’s Experiment with Pneumonia
and the accidental discovery of
Transformation
•Frederick Griffiths was a
bacteriologist studying
pneumonia
•He discovered two types
of bacteria:
–Smooth colonies
–Rough colonies
:

3. Griffith’s Experiment with Pneumonia
and the accidental discovery of
Transformation
•When heat was
applied to the deadly
smooth type…
•And injected into a
mouse…
•The mouse lived!

4. •Griffith injected the heat-killed
type and the non-deadly rough
type of bacteria.
•The bacteria “transformed”
itself from the heated non-
deadly type to the deadly type.
Griffith’s Experiment with Pneumonia
and the accidental discovery of
Transformation

5. Griffith’s Experiment did not prove
that DNA was responsible for
transformation
How would you design an
experiment to prove that DNA
was responsible for
transformation?

6. Avery, McCarty, and MacLeod
Added the non-deadly Rough Type of
Bacteria to the Heat-Killed Smooth Type
CarbohydratesCarbohydratesLipidLipid
ss
ProteinsProteins RNARNA DNADNA
To the Heat-Killed Smooth Type, To the Heat-Killed Smooth Type,
added enzymes that destroyed…added enzymes that destroyed…

7. S-Type S-Type
Carbohydrates Carbohydrates
DestroyedDestroyed
S-Type S-Type
Lipids Lipids
DestroyeDestroye
dd
S-Type S-Type
Proteins Proteins
DestroyeDestroye
dd
S-Type S-Type
RNA RNA
DestroyedDestroyed
S-Type S-Type
DNA DNA
DestroyedDestroyed
Conclusion:Conclusion:
DNA was the DNA was the
transforming factor!transforming factor!

8. The Hershey-Chase Experiment
Alfred Hershey &
Martha Chase
worked with a
bacteriophage:
A virus that
invades bacteria.
It consists of a
DNA core and a
protein coat
DNADNA
Protein coatProtein coat

9. Protein coats of bacteriophages labeled with Sulfur-Protein coats of bacteriophages labeled with Sulfur-
3535
DNA of bacteriophages labeled with Phosphorus-32DNA of bacteriophages labeled with Phosphorus-32
BacteriumBacterium
BacteriumBacterium
PhagePhage
PhagePhage
1.1.Hershey and Chase Hershey and Chase
mixed the mixed the
radioactively-labeled radioactively-labeled
viruses with the viruses with the
bacteriabacteria
The viruses infect The viruses infect
the bacterial cells.the bacterial cells.

10. Protein coats of bacteriophages labeled with Sulfur-Protein coats of bacteriophages labeled with Sulfur-
3535
DNA of bacteriophages labeled with Phosphorus-32DNA of bacteriophages labeled with Phosphorus-32
2.2.Separated the Separated the
viruses from the viruses from the
bacteria by agitating bacteria by agitating
the virus-bacteria the virus-bacteria
mixture in a blendermixture in a blender

11. Protein coats of bacteriophages labeled with Sulfur-Protein coats of bacteriophages labeled with Sulfur-
3535
DNA of bacteriophages labeled with Phosphorus-32DNA of bacteriophages labeled with Phosphorus-32
3.3.Centrifuged the mixture so that Centrifuged the mixture so that
the bacteria would form a pellet the bacteria would form a pellet
at the bottom of the test tubeat the bottom of the test tube
4.4.Measured the radioactivity in the Measured the radioactivity in the
pellet and in the liquidpellet and in the liquid

12. How does DNA replicate?
Hypotheses:
ConservativeConservative Semi-Semi-
ConservativeConservative
DispersiveDispersive

13. 1.1.Bacteria cultured in medium Bacteria cultured in medium
containing a containing a heavyheavy isotope of isotope of
Nitrogen (Nitrogen (
1515
N)N)
Meselson-Stahl ExperimentMeselson-Stahl Experiment

14. 2.2.Bacteria transferred to a medium Bacteria transferred to a medium
containing elemental Nitrogen (containing elemental Nitrogen (
1414
N)N)
Meselson-Stahl ExperimentMeselson-Stahl Experiment

15. Meselson-Stahl ExperimentMeselson-Stahl Experiment
3.3.DNA sample centrifuged after First DNA sample centrifuged after First
replicationreplication

16. Meselson-Stahl ExperimentMeselson-Stahl Experiment
4.4.DNA sample centrifuged after DNA sample centrifuged after
Second replicationSecond replication

17. DNA replication
E.Coli DNA polymerase I requires:E.Coli DNA polymerase I requires:
1. All four dNTPs (dATP, dGTP, dCTP and dTTP)1. All four dNTPs (dATP, dGTP, dCTP and dTTP)
2. A primer chain with a free 3`-OH end2. A primer chain with a free 3`-OH end
3. A template strand to which the primer is 3. A template strand to which the primer is
basepairedbasepaired
• • Double-stranded DNA that is fully intact andDouble-stranded DNA that is fully intact and
lacking a free 3`-OH end will not be replicatedlacking a free 3`-OH end will not be replicated
(Ex: Intact circular DNA)(Ex: Intact circular DNA)
4. Mg2+4. Mg2+

18. DNA synthesis: DNA Polymerase Reaction
(DNA)n + dNTP (DNA)n+1 + PPi 2Pi(DNA)n + dNTP (DNA)n+1 + PPi 2Pi
PrimerPrimer 5` n+1 3`
→→
5` n+1 3`
→→
5` n+2 3`
→
5` n+2 3`
→
TemplateTemplate
DNA chain growth is 5’ to 3’DNA chain growth is 5’ to 3’

19. Summary of basic mechanism of DNA replicationSummary of basic mechanism of DNA replication
• Replication is semiconservative
• DNA polymerase requires a template-primer
complex
• dNTPs are the substrates for DNA synthesis
• PPi breakdown to 2 Pi (catalyzed by
pyrophosphatase) drives DNA synthesis
• DNA Polymerase accuracy: 1 mistake every 108
bases