DNA
The Genetic Material
AP Biology
Scientific History
 The march to understanding that DNA is
the genetic material
T.H. Morgan (1908)
 Frederick Griffith (1928)
 Avery, McCarty & MacLeod (1944)
 Erwin Chargaff (1947)
 Hershey & Chase (1952)
 Watson & Crick (1953)
 Meselson & Stahl (1958)

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1908 | 1933
Chromosomes related to phenotype
 T.H. Morgan

working with Drosophila
 fruit flies

associated phenotype with
specific chromosome
 white-eyed male had specific
X chromosome
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1908 | 1933
Genes are on chromosomes
 Morgan’s conclusions
________________________
 but is it the protein or the
DNA of the chromosomes
that are the genes?

 initially proteins were thought
to be genetic material…
Why?
What’s so impressive
about proteins?!
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The “Transforming Principle”
 Frederick Griffith

Streptococcus pneumonia bacteria
 was working to find cure for pneumonia
harmless live bacteria (“rough”)
mixed with heat-killed pathogenic
bacteria (“smooth”) causes fatal
disease in mice
 a substance passed from dead
bacteria to live bacteria to change
their phenotype


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_____________________________
1928
The “Transforming Principle” mix heat-killed
live pathogenic
strain of bacteria
A.
mice die
live non-pathogenic heat-killed
strain of bacteria
pathogenic bacteria
B.
C.
mice live
mice live
pathogenic &
non-pathogenic
bacteria
D.
mice die
Transformation = change in phenotype
something in heat-killed bacteria could still transmit
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disease-causing properties
1944
DNA is the “Transforming Principle”
 Avery, McCarty & MacLeod

purified both DNA & proteins separately from
Streptococcus pneumonia bacteria
 which will transform non-pathogenic bacteria?

injected protein into bacteria
 no effect

injected DNA into bacteria
 transformed harmless bacteria into
virulent bacteria
mice die
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What’s the
conclusion?
1944 | ??!!
Avery, McCarty & MacLeod
 Conclusion

___________________________________________
___________________________________________
Oswald Avery
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Maclyn McCarty
Colin MacLeod
1952 | 1969
Confirmation of DNA
 Hershey & Chase
classic “blender” experiment
 worked with bacteriophage

 viruses that infect bacteria

Why use
Sulfur
vs.
Phosphorus?

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grew phage viruses in 2 media,
radioactively labeled with either
 ____________________
 ____________________
infected bacteria with
labeled phages
Hershey
Protein coat labeled
with 35S
Hershey
& Chase
DNA labeled with 32P
T2 bacteriophages
are labeled with
radioactive isotopes
S vs. P
bacteriophages infect
bacterial cells
bacterial cells are agitated
to remove viral protein coats
Which
radioactive
marker is found
inside the cell?
Which molecule
carries viral
genetic
info?
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35S
radioactivity
found in the medium
32P
radioactivity found
in the bacterial cells
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Blender experiment
 Radioactive phage & bacteria in blender

___________________
 radioactive proteins stayed in supernatant
 therefore viral protein did NOT enter bacteria

___________________
 radioactive DNA stayed in pellet
 therefore viral DNA did enter bacteria

___________________________________
Taaa-Daaa!
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1952 | 1969
Hershey
Hershey & Chase
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Martha Chase
Alfred Hershey
1947
Chargaff
 DNA composition: “________________”
varies from species to species
 all 4 bases not in equal quantity
 bases present in characteristic ratio

 humans:
A = 30.9%
T = 29.4%
G = 19.9%
C = 19.8%
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That’s interesting!
What do you notice?
Rules
A = T
C = G
1953 | 1962
Structure of DNA
 Watson & Crick

___________________________________
 other leading scientists working on question:
 Rosalind Franklin
 Maurice Wilkins
 Linus Pauling
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Franklin
Wilkins
Pauling
1953 article in Nature
Watson and Crick
Watson
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Crick
Rosalind Franklin (1920-1958)
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But how is DNA copied?
 Replication of DNA

base pairing suggests
that it will allow each
side to serve as a
template for a new
strand
“It has not escaped our notice that the specific pairing we have postulated
immediately suggests a possible copying mechanism for the genetic
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material.”
— Watson & Crick
Models of DNA Replication
 Alternative models

become experimental predictions
semiconservative
P
1
2
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Can you design
a nifty experiment
to verify?
semiconservative
semiconservative
Semiconservative replication
1958
 Meselson & Stahl


label “parent” nucleotides in DNA strands with
heavy nitrogen = 15N
label new nucleotides with lighter isotope = 14N
“The Most Beautiful Experiment in Biology”
Make predictions…
15N/15N
15N
parent
strands
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parent
replication
Predictions
________
1st round of
replication
________
________
________
semiconservative
dispersive
conservative
2nd round of
replication
________
P
________
________
________
________
1
15N/15N
2 15N parent
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strands
semiconservative
dispersive
conservative
Meselson & Stahl
Matthew Meselson
Franklin Stahl
Franklin Stahl
Matthew Meselson
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Scientific History
 March to understanding that DNA is the genetic material

T.H. Morgan (1908)
 ___________________________________________________

Frederick Griffith (1928)
 ___________________________________________________

Avery, McCarty & MacLeod (1944)
 ___________________________________________________

Erwin Chargaff (1947)
 ___________________________________________________

Hershey & Chase (1952)
 ___________________________________________________

Watson & Crick (1953)
 ___________________________________________________

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Meselson & Stahl (1958)
 ___________________________________________________
The “Central Dogma”
 Flow of genetic information in a cell
translation
DNA
replication
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translation
RNA
protein
Science …. Fun
Party Time!
Any Questions??
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DNA Replication
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Watson and Crick
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1953 article in Nature
Double helix structure of DNA
“It has not escaped our notice that the specific pairing we have postulated
immediately suggests a possible copying mechanism for the genetic
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material.”
Watson & Crick
Directionality of DNA
 You need to
PO4
nucleotide
number the
carbons!

it matters!
N base
CH2
This will be
IMPORTANT!!
O
ribose
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OH
The DNA backbone
 Putting the DNA
backbone together

refer to the 3 and 5
ends of the DNA
 the last trailing carbon
Sounds trivial, but…
this will be
IMPORTANT!!
PO4
base
5 CH2
O
4
1
C
3
O
–O P O
O
5 CH2
2
base
O
4
1
3
OH
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2
Anti-parallel strands
 Nucleotides in DNA
backbone are bonded from
phosphate to sugar
between 3 & 5 carbons
DNA molecule has
“direction”
 complementary strand runs
in opposite direction

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Bonding in DNA
5
hydrogen
bonds
3
covalent
phosphodiester
bonds
3
5
….strong or weak bonds?
AP
Biology
How
do the bonds fit the mechanism for copying DNA?
Base pairing in DNA
 ________________
______________
 ______________

 ________________
______________
 ______________

 Pairing

______________
 2 bonds

______________
 3 bonds
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Copying DNA
 Replication of DNA
base pairing allows
each strand to serve as
a template for a new
strand
 new strand is 1/2
parent template &
1/2 new DNA

 ____________________
copy process
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DNA Replication
Let’s meet
the team…
 Large team of enzymes coordinates replication
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Replication: 1st step
 Unwind DNA

I’d love to be
helicase & unzip
your genes…
__________________
 unwinds part of DNA helix
 stabilized by _____________________________
helicase
single-stranded binding proteins
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replication fork
Replication: 2nd step
 Build daughter DNA
strand
add new
complementary bases
 ___________________

DNA
Polymerase III
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But…
Where’s the
We’re missing
ENERGY
something!
for the bonding!
What?
Energy of Replication
Where does energy for bonding usually come from?
We come
with our own
energy!
You
remember
ATP!
Are there
other ways
to get energy
out of it?
energy
GTP
TTP
ATP
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modified nucleotide
And we
leave behind a
nucleotide!
TMP
GMP
AMP
ADP
Energy of Replication
 The nucleotides arrive as nucleosides

DNA bases with P–P–P
 P-P-P = energy for bonding


DNA bases arrive with their own energy source
for bonding
bonded by enzyme: ________________________
ATP
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GTP
TTP
CTP
5
Replication
 Adding bases

3
energy
DNA
Polymerase III
can only add
nucleotides to
3 end of the
growing DNA strand
 need a primer
nucleotide to
bond to

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_________________
B.Y.O. ENERGY!
The energy rules
the process
3
5
5
3
no energy
to bond
AP Biology
3
5
3

5
3
5
5
3
5
3
ligase
energy
AP Biology
3
5
3
5
Okazaki
Leading & Lagging strands
Limits of DNA polymerase III

can only build onto 3 end of
an existing DNA strand
5
3
5
3
5
3
5
5
5
Lagging strand
ligase
growing
3
replication fork
Leading strand
3
___________________


__________________
__________________
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 “spot

3
welder” enzyme

5
3
DNA polymerase III
__________________

continuous synthesis
Replication fork / Replication bubble
3
5
5
3
DNA polymerase III
leading strand
5
3
3
5
3
5
5
5
3
lagging strand
3
5
3
5
lagging strand
5
5
leading strand
3
growing
replication fork
leading strand
3
lagging strand
5 5
AP Biology
growing
replication fork 5
5
5
3
Starting DNA synthesis: RNA primers
Limits of DNA polymerase III

can only build onto 3 end of
an existing DNA strand
5
3
3
5
5
3
5
3
5
growing
3
replication fork
DNA polymerase III
primase
RNA 5
______________________
built by ________________
 serves as starter sequence
DNA polymerase III
AP for
Biology

3
Replacing RNA primers with DNA
______________________

removes sections of RNA
primer and replaces with
DNA nucleotides
DNA polymerase I
3
5
5
5
3
ligase
growing
3
replication fork
RNA
5
3
But DNA polymerase I still
can only build onto 3 end of
an
existing DNA strand
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Chromosome erosion
DNA polymerases can
only add to 3 end of
an existing DNA strand
Houston, we
have a problem!
DNA polymerase I
5
3
3
5
5
growing
3
replication fork
DNA polymerase III
5
Loss of bases at 5 ends
in every replication
chromosomes get shorter with each replication
AP
Biologyto number of cell divisions?
 limit

3
Telomeres
Repeating, non-coding sequences at the end
of chromosomes = protective cap

limit to ~50 cell divisions
5
3
3
5
growing
3
replication fork
5
____________________



enzyme extends telomeres
can add DNA bases at 5 end
different level of activity in different cells
AP Biology
 high in stem cells & cancers -- Why?
telomerase
5
TTAAGGG TTAAGGG 3
Replication fork
3’
5’
5’
3’
5’
3’
5’
3’
direction of replication
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DNA polymerases
 DNA polymerase III
1000 bases/second!
 main DNA builder

Roger Kornberg
2006
 DNA polymerase I
20 bases/second
 editing, repair & primer removal

DNA polymerase III
enzyme
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Arthur Kornberg
1959
Editing & proofreading DNA
 1000 bases/second =
lots of typos!
 DNA polymerase I

proofreads & corrects
typos

repairs mismatched bases

removes abnormal bases
 repairs damage
throughout life

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reduces error rate from
1 in 10,000 to
1 in 100 million bases
Fast & accurate!
 It takes E. coli <1 hour to copy
5 million base pairs in its single
chromosome

divide to form 2 identical daughter cells
 Human cell copies its 6 billion bases &
divide into daughter cells in only few hours
remarkably accurate
 only ~1 error per 100 million bases
 ~30 errors per cell cycle

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What does it really look like?
1
2
3
4
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Any Questions??
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