Discovery and Replication of

DNA
Genes are physical things carried
by chromosomes
If genes are physical, they must be made
of molecules that can be purified.

How to purify something:

1) Separate components of a complex mixture.

2) Assay each component to determine which

has the desired property
 Assays
• An “assay” is just a way of measuring
something.
• It can measure a substance, like the assay
for starch.
• It can also measure an abstract
phenomenon, like an assay for memory
(i.e. mice in a maze).
 How to purify DNA

• Grind up your favorite organism.

• Extract lipids and proteins with organic
solvent (e.g. phenol).
• Precipitate with ethanol.
Friedrich Miescher
 But how to assay genetic
materialness (materialhood?) ?
A hypothetical assay for genetic material
How to purify genetic material:
Fredrick Griffith and the transforming principle
Oswald Avery uses Griffith’s assay to
identify the transforming principle

Substance Converts R to S?

Lipid no

Carbohydrate no

Protein no

DNA YES
Does DNA work in other
organisms?
Yes, it even works in viruses
 DNA
Protein

Bacterium
 The phage injects its
genetic material into
the bacterium

The genetic material

directs the generation of
more phage

The new phage burst

out to infect new
bacteria
 The Hershey-Chase Experiment

You can specifically label protein with

radioactive sulphur and label DNA with radioactive
phosphate
DNA is the Transforming
Principle

Also known as the genetic material

Clues to the mystery of DNA structure:

1) Chemical structure of monomers

2) Chargaff’s rules
3) Crystallography
 Chargaff’s Rules:

A = T and G = C

The ratio A + T / G + C varies with

organism
Rosalind Franklin, Maurice Wilkins
 Crystalography
• DNA consists of two strands.
• These strands twist around each other in a
double helix.
• The phosphates are probably on the outside.
• The strands run antiparallel to each other.

5’ 3’

3’ 5’
Watson and Crick put it all together
DNA strands are the reverse
complement of each other:

Reverse because they are anti-

parallel (read in opposite
directions going 5’ to 3’)

Complement because A on one

strand corresponds to T on the
other strand, G to C etc.
Complementary match …compared to non-optimal
optimizes number of match. Note purine must
hydrogen bonds… pair with pyrimidine.
CH2
H2N
O
T C
N N N
HN
N N N
N O
A O A
N N H
N H N

CH2
H2N
O
C T
N N N
HN
N NH N NH
O O
G G
N NH2 N NH2
N N
 RNA can also
form a
complementary
strand with DNA.
 The key to DNA:
• The sequence of nucleotides is not constrained
by the structure. DNA can accommodate any
sequence. An arbitrary sequence can encode
information

• The two strands of DNA encode the same

information in complementary form suggesting a
method of replication.
How does DNA make copies of
itself?
Replication
Models

Newly
synthesized
strands
Centrifuging DNA in a tube
of cesium chloride
separates heavy 15N from
light 14N DNA

Heavy 15N DNA sinks faster

and ends up closer to the
bottom

Light 14N DNA sinks slower

and remains closer to the
top of the tube.
Meselson and Stahl experiment
To make DNA in a test tube just
add:

• Triphosphate nucleotides.
• DNA polymerase.
• Template DNA with...
• Ragged ends.
 Problems:

• How to pry strands apart.

• How to make ragged ends.

Solutions:
• The enzyme helicase unwinds the
DNA strands.
• The enzyme primase makes short
RNA primers
Origin or Replication
Replication Fork

Replication
Bubble
primase
Replication proceeds
 One origin of Many origins
replication of replication

Replication
spreads in both Replication
directions from spreads in both
origin until forks directions from
moving in origin until forks
opposite moving in
directions meet opposite
at the opposite directions meet
side of the circle in the middle
(ter)
Next Lecture: the genetic code