9 | DNA STRUCTURE AND ANALYSIS

DNA

serves as the genetic material except in viruses

right handed double helix (James-Crick model)
Strands are antiparallel and are held together through hydrogen bonds:
o A - -T (2 H bonds)
o G - - - C ( 3 H bonds)
structure is for storing and expressing the genetic information

RNA

single stranded
serves as the genetic material in some viruses

Chromosomes

structures that contain genes controlling the phenotypic traits that are transmitted through
gametes offspring
contains both a nucleic acid and a protein component

Genes

influences the form and characteristics of each individual

o genetic material

9.1 | THE GENETIC MATERIAL MUST EXHIBIT FOUR CHARACTERISTICS

1. Replication
Fundamental property of all living organisms
Doubling the amount of the genetic material
o Must be equally partitioned into the daughter cells
Formation of the gametes also replicates and partition the genetic material, only getting of
the original genetic materialMeiosis
2. Storage of information
molecule has to act as a repository of genetic information that may or may not be expressed by
the cell in which it resides.
a. Ex. Turn on and off in bacteria and higher animals
i. Bacteria specific environmental conditions
ii. Human melanin genes
3. Expression of information
basis of the process of information flow within the cell

Transcription of DNA
o 3 RNA molecule are synthesized: mRNA, rRNA and tRNA
o mRNA- translated into proteins; product of a specific gene and directs the synthesis of a
different protein
Transltion
o
chemical information in mRNA directs the construction of a chain of amino acids called
polypeptides, which fold into a protein.

Variation by mutation

Mutation
o a change in the chemical composition of DNA;
alteration is reflected during transcription and
translation that affects a specific protein
o can be passed to the future generations
genetic
variation
also
include
the
alterations
of
chromosome number and rearrangements within and
between chromosomes providing a new raw material for
the process of evolution

9.2
Genetic Material

| Until 1994, Observations Favored Protein as the

Since there is a great diversity of proteins in abundant cells

Friedrich Miescher

Isolated cell nuclei and derived an acid substance containing DNA called nuclein
DNA lacks diversity necessary to store extensive genetic information
The conclusion was based on Phoebus A Levene
o 1910; DNA contained approximately equal amounts of 4 similar molecules called nucleotides.
o Tetranucleotide hypothesis

Erwann Chargaff

Showed that Levene was incorrect

Most organisms do not contain precisely equal proportions of the 4 nucleotides

9.3 | Evidence Favoring DNA as the Genetic Material was First Obtained During the
Study of Bacteria and Bacteriophages

Oswald Avery, Colin MacLeod and Maclyn McCarty published

principle
DNA was carrier of the genetic information
DNA was the responsible on in heredity

the chemical nature of a transforming

TRANSFORMATION: EARLY STUDIES

Frederick Griffifth

Where the work of Avery, MacLeod and MacCarteys was initianted in 1927
Experimented in Diploccocus pneumonia
o Some are virulent strains and some are avirulent strains
Virulent strains
o Have the polysaccharide capsule
o Are not engulfed easily ; multiply and causes oneumoniae
o Form smooth colonies (S) with shiny surface on an agar culture plate
Avirulent
o do not have the polysaccharide capsule
o Readily engulf and destroyed by the phagocytic cells in the animals circulatory system
o
Produces rough colonies (R )
o If heated, did not cause pneumonia
Diplococcus may be one of dozens of different types called serotypes
o Specificity is due to the detailed chemical structure of the polysaccharide constituent of
the thick, slimy capsule.

Heated IIIS (virulent) + living IIR( avirulent)= still caused pneumonia

o The possibility that the avirulent bacteria has changed or mutated the IIS in the absence
of the heat-killed IIIS bacteria
o An interaction took place between the IIR and heat killed IIIS cells
o Transformation
Converted avirulent cells into virulent IIIS cells.
Transforming principle might be some part of the polysaccharide capsule or compound
required for capsule synthesis, although capsule alone did not cause pneumonia.
The dead IIS served as a pabulum for the IIR cells

Henry Dawson

Showed that transformation could occur in vitro tube containing only bacterial cells.
Injection to the mice is not necessary for transformation to occur.

Lionel J. Alloway

Refined the in vitro experiments using extracts from S cells added to living R cells
The soluble filtrate from the heat-killed IIIS cells was as effective inducing transformation as were the
intact cells.
Did not view transformation as a genetic event but a physiological modification

TRANSFORMATION: Avery, MacLeod and McCarty Experiment

Obtained the transforming principle in a highly purified state DNA

Check Figure 3 for summary

Soluble filtrate -derived(retained the ability to

induce transformation of type IIR avirulen cells

Preotease- protein-digesting enzyme

Ribonuclease- RNA digesting enzyme

But when deoxyribonuclease was used, the

transforming activity present in the filtrate was
destroyed.

Conclusion in 1944 publication:

The evidence presented supports the belied that a nucleic
acid of deoxyribose type is the fundamental unit of the
transforming orinciple of Pneumococcu type III
Transforming principles interacts with the IIR cell and
gives rise to a coordinated enzymatic reactions that culminates in the synthesis of the type IIIS capsular
polysaccharide

Transformation is heritable and the process affects the genetic material.

THE HERSHEY-CHASE EXPERIMENT

E. coli and a bacteriophage T2

Phage or bacteriophage
o
virus consists of a protein coat surrounding a core of DNA.
o External structure is composed of a hexagonal head plus a tail
o Adsorbs to the bacterial cell and some component of the phage enters the cell.
o Viral information commandeers- responsible for the viral reproduction manipulating the cellular
machinery of the host

1. T2 phages consists of approximately

50% protein
and 50% DNA
2. Infection is initiated by adsorption of the phage by its tail fibers to the bacterial cell
3. The production if new virus occurs within the bacterial cell.

Used radioisotopes to follow the molecular components of phages during infection

32
P labels DNA
o DNA contains phosphorus but not sulfur

35

S labels protein
Proteins contain sulfur bot not phosphorus

E.coli cells are first grown in the presence of either radioactive isotope and the infected with T2 virus, the
progeny ohage will then be labeled with a DNA core or a labeld Protein coat.
The radioactive phages can be isolated and infect unlabeled bacteria
1

labeled phage and unlabeled bacteria were mixed

adsorption complex was formed as the phages attached

their tail fibers to the bacterial wall
3 comlexes were isolated and subjected to shear forceusing a blender
4 force stripped off the attached phages; analyzed
separately
5 radioisotopes were traced
6
7

P DNA - bacterial cell

S protein- remaIned outside. "ghosts"
bacterial cells were lysed as new phages were produced,
the progeny contained 32P but not 35S
32
35

Protein of the coat remains outside the host cell and is not involved in the production of new phages
DNA enters the host cell and directs the phage reproduction

Genetic material in phage T2 is DNA, and not protein

TRANSFECTION EXPERIMENTS

1957, E. coli was treated with the enzyme lysozyme, the outer wall of the cell can be removed
without destroying the bacterium.
o Making them naked, containing only the cell membranes as the outer boundary of cell
o Structures are called as protoplasts or spheroplasts.

John Spizizen and Dean Fraser

o Used protoplasts and were able to initiate the phage multiplication with distrupted T2 particles.
o A virus did not have to be intact for infection to occur

Transfection
o Process of infection by only the viral acid
o Used only purified DNAfrom bacteriophages

DNA serves as the genetic material in all organism

9.4 | Indirect and Direct Evidence Supports the Concept that DNA Is the Genetic
Material in Eukaryotes
Indirect Evidence: Distribution of DNA

The genetic material should be found where it functions

o Nucleus as part of the chromosomes
DNA and protein
But proteins are also abundant in the cutoplasm
Mitochondria and chloroplasts both perform genetic functions; DNA is present
DNA is found only where primary genetic function is known to occur

Correlation between the ploidy(,n, 2n etc.) of cells and the quantity of the molecule
that functions as the genetic material occurs.

No consistent correlation occurs between gametes and diploid cells for proteins

Indirect Evidence: Mutagenesis

Ultrviolet (UV) light

One of the agents capable od inducing mutations in the genetic material

o Action spectrum
Obtained when data are plotted
UV light as mutagenic agent is obtained
o Absorption spectrum
Where action spectrum is compared
Of any molecule suspected to be genetic material

The molecule serving as the genetic material is expected to absorb at the wavelengths
shown to be mutagenic.

Uv light is most mutagenic at the wavelength () of about 260 nm

Both DNA and RNA absorb UV light at 260 nm
Proteins absorb at around 280 nm but no mutagenic effects are observed

Direct Evidence: Recombinant DNA studies

Segments of eukaryotic DNA corresponding to

specific genes are isolated and spliced into
bacterial DNA

Complex can be inserterted into a bacterial

cell
genetic expression is monitored

introduction of eukaryotic gene

presence of corresponding eukaryotic

preotein product demonstrates directly the
DNA is functional in the bacterial cell

As the bacterium divides, the eukaryotic DNA replicates along the host DNA

Beatrice Mintz

Demonstrated the DNA encoding the human B-globin protein

Microinjected to a fertilized mouse egg

o
o

Later found to be present and expressed in adult mouse tissue and it is transmitted to and expressed
in the mouses progeny
The mouses are called to be TRANSGENIC ANIMALS

9.5 | RNA serves as the Genetic Material in Some Virus

Virus core= RNA

Tobacco mosaic virus (TMV)
o Spread on its leaves
o Lesions were caused by the viral infection

Norman Pace and Sol Spiegelman

RNA from phage QB can be isolated and replacted in vitro.

RNA replicase
o Enzyme that causes replication

Retrovirus
o Replicate in an unusual way
o RNA serves as a template for synthesis of the complementary DNA molecule
o Process is called as the reverse transcription
Reverse transcriptase
DNA polymerase that direct the direction of RNA- dependent DNA polymerase
The DNA intermediate can be incorporated into the genome of the host cell
DNA transcription- copies of the retroviral RNA chromosome are produced
Includes HIV- human immunodeficiency virus
o That causes aids and RNA tumor virus