DNA: The Genetic Material
Dr. Henry O. Ogedegbe
Department of EHMCS
The Hammerling Experiment: Cells Store
Hereditary Information in the Nucleus
• Where is hereditary information stored in the cell?
• The Danish biologist Hammerling cut cells into pieces to
see which were able to express hereditary information
• He chose the green alga Acetabularia which grows up to 5
cm as a model organism for his experiments
• The genus Acetabularia have distinct foot, stalk and cap
regions and the nucleus is located in the foot
• He amputated the stalk of some cells and the feet of others
• He found that when he amputated the cap, a new cap
regenerated from the remaining portions of the cell
The Hammerling Experiment: Cells Store
Hereditary Information in the Nucleus
• When the foot was amputated however, no new foot
regenerated from the cap or the stalk
• He therefore hypothesized that the hereditary information
resided within the foot of Acetabularia
• His hypothesis was tested by selecting individuals from
two species of the genius Acetabularia which had different
caps:
– A. mediterranea has a disc shaped cap
– A. crenulata has a branched flower-like cap
• He grafted a stalk from A. crenulata to a foot from A
mediterranea
The Hammerling Experiment: Cells Store
Hereditary Information in the Nucleus
• The cap regenerated looked somewhat like the cap of A.
crenulata
• He then cut the regenerated stalk and all subsequent caps
were disc shaped like like A. mediterranea
• The experiment supported Hammerling’s hypothesis
Transplantation Experiments: each Cell
Contains s Full Set of Genetic Instructions
• Robert Briggs and Thomas King tested the hypothesis that
that the nucleus is the repository of hereditary information
• They removed the nucleus from frog eggs and found out
that without the nucleus, the egg did not develop
• When the nucleus was replaced, with one from a frog
embryo cell, the egg developed into an adult frog
• While this experiment produced abnormal frogs
modifications of the experiment by other workers
produced satisfactory results
The Griffith Experiment: hereditary
Information Can Pass between organisms
• Discovery of Transformation:
• Griffith performed experiments in which he injected mice
with virulent strain of streptococcus pneumoniae
• The infected mice all died.
• When he infected similar mice with mutant strains of S.
pneumoniae that lacked the virulence factor the mice
showed no ill effect
• When he infected mice with dead mutant strain of S.
pneumoniae the mice remained healthy
• Similarly, infection of the mice with the R form of the
bacteria produced no ill effect in the mice
The Griffith Experiment: hereditary
Information Can Pass between organisms
• When he infected similar mice with a mixture of dead
virulent bacteria and the R form some of the mice died
• The virulence factor had been transferred to the R form
which transformed the coatless form to the virulent form
The Avery Experiment: The
Transforming Principle Is DNA
• Avery and co-workers characterized the transforming
principle
• They prepared a mixture of dead and coatless S.
pneumoniae similar to what Griffith had done
• Then they removed all the proteins from the mixture
• Despite the removal of the proteins, the transforming
activity of the mixture was not reduced
• The properties of the transforming principle resembled
those of DNA in many respects
The Avery Experiment: The
Transforming Principle Is DNA
• Analysis of the purified principle produced elements which
agreed closely with DNA
• In an ultracentrifuge the transforming principle migrated
like DNA
• Removal of lipids and proteins from the principle did not
diminish its activity
• Protein digesting enzymes did not affect the principle
• The DNA digesting enzyme DNase destroyed all the
transforming principle
The Hershey-Chase Experiment: Some
Viruses Direct Their Heredity with DNA
• Hershey-Chase experiment involved bacteriophages,
viruses that attack bacteria
• They employed the bacteriophage T2 which is a DNA
virus
• They labeled the viral DNA with radioactive isotope of
phosphorus 32P and the protein coat with radioactive sulfur
35S
• After the labeled viruses were allowed to infect the
bacteria the bacterial cells were agitated violently
• This was designed to remove the protein coats of the
infecting viruses from the surface of the bacteria
The Hershey-Chase Experiment: Some
Viruses Direct Their Heredity with DNA
• The 32P label had transferred to the interior of the bacteria
and viruses released subsequently contained the 32P label
• The hereditary information injected into the bacteria that
specified new generation of viruses was DNA
• Thus the DNA is clearly the repository of hereditary
information
The Frankel-Conrat Experiment: Other
Viruses Direct Their Heredity with RNA
• Fraenkel-Conrat experimented with RNA viruses to
determine how they reproduce
• They employed the tobacco mosaic virus and the Holmes
ribgrass virus.
• They separated the RNA from the proteins and discovered
that the RNA molecules were still infective whereas the
protein molecules were not
The Chemical Nature of Nucleic Acid
• The DNA was discovered in 1969 by Friedrich Miescher
four years after the publication of Mendel’s work
• He extracted a white substance from human cells and fish
sperm
• The proportion of nitrogen and phosphorus in the
substance was different from any previous substances
• This convinced him that he was dealing with a new
substance
• Due to its slight acidity, it came to be known as nucleic
acid
The Chemical Nature of Nucleic Acid
• The primary structure was elucidated in the 1920s by the
biochemist P.A. Levene
• DNA contains three components which include the
phosphate group, five carbon sugars, and nitrogenous
bases
• The nitrogenous bases are purines; adenine guanine and
pyramidines; thymine, and cytosine
• RNA contains uracil instead of thymine
• A nucleotide consist of a sugar attached to a phosphate
group and a base
The Chemical Nature of Nucleic Acid
• The four carbon atoms and the oxygen atom form a five
membered ring
• The carbon atoms are numbered 1’ to 5’ proceeding
clockwise from the oxygen atom
• The prime symbol indicates that the carbon refers to a
carbon in a sugar rather than a base
• The subunits are linked together by phosphodiester bonds
• The resultant two-unit polymer still has a free 5’ phosphate
group at one end and a free 3’ hydroxyl group at the other
end
The Chemical Nature of Nucleic Acid
• Chargaff’s Analysis showed that the nucleotide
composition of DNA molecules varies in complex ways
• This led to Chargaff’s rules:
– The proportion of A always equals that of T and the proportion of
G always equals that of C
– There is always an equal proportion of purines (A and G) and
pyramidines (C and T)
The three-Dimensional Structure of DNA
• The work of Rosalind Franklin involved X-ray
crystallographic analysis of DNA
• This involved bombarding the DNA molecules with beams
of X-rays
• Rosalind used DNA in the form of fibers in the laboratory
of Maurice Wilkins
• The work of Rosalind led to the discovery of the double
helix by Crick and Watson
• The double helix is stabilized by antiparallel strands one
chain running 3’ to 5’ the other 5’ to 3’
The Meselson-Stahl Experiment: DNA
Replication Is semiconservative
• The basis for copying the genetic information is
complementarity
• If the DNA molecule is unzipped one would need only to
assemble the appropriate complementary nucleotides
• This would produce two daughter duplexes with the same
sequence
• This form of DNA replication is called semiconservative
because the sequence of the original duplex is conserved
• Each strand of the duplex becomes part of another duplex
The Replication Complex
• The DNA polymerase III plays a very essential part in gene
DNA replication
• The polymerase III is a complex of 10 different kinds of
polypeptide chains
• The enzyme is a dimer with two similar multisubunit
complexes
• Polymerase III threads the DNA through the complex at
the rate of 1000 nucleotides per second
The Replication Complex
• The two strand of DNA are assembled differently
• The polymerase III can add nucleotide only to the 3’ end of
a DNA strand
• That means that replication occurs in the 5’ to 3’ direction
on a growing DNA strand
• The leading strand is built up by adding nucleotides
continuously to it growing 3’ end
• The lagging strand which elongates away from the
replication fork is synthesized discontinuously as short
segments
The Replication Complex
• These discontinuous segments are called the Okasaki
fragments
• They are about 100 to 200 nucleotides long in eukaryotes
and about 1000 to 2000 nucleotides long in prokaryotes
• The Okasaki fragment is synthesized by DNA polymerase
III in the 5’ to 3’ direction
• The overall replication of the DNA is said to be
semidiscontinuous
The Replication Process
• The replication of the DNA molecule takes place in five
interlocking steps:
– Opening of the DNA double helix
•
•
•
•
–
–
–
–
Initiation replication
Unwinding the duplex
Stabilizing the single strand
Relieving the torque generated by unwinding
Building a primer
Assembling complementary strands
Removing the primer
Joining the Okasaki fragments
The One-Gene/One-Polypeptide
Hypothesis
• The discovery that certain types of inherited diseases were
prevalent in particular families led to the conclusion that:
– These diseases were Mendelian traits which had resulted from
changes in the hereditary information in an ancestor
– An example is alkaptonuria in which patients produce urine that
contained homogentisic acid (alkapton)
– Such patients lacked the enzyme necessary to catalyze the
breakdown of alkaptonuria
• Invariably it was concluded that genes specify particular
enzymes
• This knowledge was clearly elucidated by Beadle and
Tatum in their experiments involving the bread mold
The One-Gene/One-Polypeptide
Hypothesis
• Beadle and Tatum were able to isolate mutant strains with
defective form of that enzyme
• The mutations were always located at specific chromosmal
sites and each enzyme had a different site
• Each mutant had a defect in a single enzyme caused by a
mutation at a single site on the chromosome
• They concluded that genes produce their effects by
specifying the structure of enzymes and each gene encodes
the structure of one enzyme
• This relationship was termed by them the one-gene/oneenzyme hypothesis or one-gene/one-polypeptide