Slide 1 Molecules of Genetic Inheritance

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Molecules of Genetic Inheritance



Shikha Yashveer1, Jayanti Tokas2, Shalini Jain3
and Hariom Yadav4 1Department of Molecular
Biology and Biotechnology, 2Department of
Biochemistry CCS HAU, Hisar, Haryana,
India 3Postgraduate Institute of Medical
Education and Research, Chandigarh,
India 4National Agri-Food Biotechnology
Institute, Mohali, Punjab, India Email
yadavhariom_at_gmail.com
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• The period from the early 1900s to World War II
  has been considered as the "golden age" of
  genetics
• Scientists still had not determined that DNA or
  the protein, was the hereditary material.

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• Friedrich Meischer in 1869 isolated DNA from fish
  sperm and the pus of open wounds
• Since it came from nuclei, Meischer named this
  new chemical, nuclein.
• Subsequently the name was changed to nucleic
  acid and lastly to deoxyribonucleic acid (DNA).

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• Robert Feulgen, in 1914, discovered that fuchsin
  dye stained DNA
• DNA was then found in the nucleus of
  all eukaryotic cells.

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• During the 1920s, biochemist P.A. Levene analyzed
  the components of the DNA molecule.
• He found it contained four nitrogenous bases
  cytosine, thymine, adenine, and guanine 
• deoxyribose sugar and a phosphate group
• He concluded that the basic unit (nucleotide) was
  composed of a base attached to a sugar and that
  the phosphate also attached to the sugar

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• He (unfortunately) also erroneously concluded
  that the proportions of bases were equal and that
  there was a tetra nucleotide that was the
  repeating structure of the molecule
• The nucleotide, however, remains as the
  fundamental unit (monomer) of the nucleic acid
  polymer
• There are four nucleotides those with cytosine
  (C), those with guanine (G), those with adenine
  (A), and those with thymine (T).

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• During the early 1900s, the link between Mendel's
  work and that of cell biologists resulted in the
  chromosomal theory of inheritance
• Garrod proposed the link between genes and
  "inborn errors of metabolism"
• Now the question was formed what is a gene? The
  answer came from the study of a deadly infectious
  disease pneumonia. 

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• Frederick Griffith in 1928
• Experimental proof that DNA is the genetic
  material
• Worked with 2 strains of the bacterium
  Streptococcus pneumoniae ----- R S

Gelatinous capsule

• Makes colony large
• Smooth appearance
• Protects the bacterium from defense mechanism
  of infected animal
• Virulent
• Causes pneumonia

• Capsular polysaccharide absent
• Small colonies
• Rough surface
• Avirulent

S
R
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Died
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Mouse remains healthy
Mouse remains healthy
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Mice died of pneumonia
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SOME SUBSTANCE HAS TRANSFORMED R TYPE OF
BACTERIA INTO S TYPE
Transforming principle
Chemical nature not known
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Oswald Avery, Colin Macleod Maclyn McCarty
(1944) R cells S cells
DNA
TRANSFORMATION
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R colonies few S colonies
Culture of R cells
Transforming activity Not Protein or RNA
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R colonies only
Culture of R cells
S cells extract
Transforming activity most likely DNA
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• Their evidence was strong but not totally
  conclusive
• The then-current favorite for the hereditary
  material was protein DNA was not considered by
  many scientists to be a strong candidate.

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• The breakthrough in the quest to determine the
  hereditary material came from the work of Max
  Delbruck and Salvador Luria with Bacteriophages
  in the 1940s
• Bacteriophages  are a type of virus that attacks
  bacteria
• The viruses that Delbruck and Luria worked with
  were those attacking Escherichia coli
• Bacteriophages consist of protein coats covering
  DNA making them ideal to resolve the nature of
  the hereditary material.

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DNA has phosphorous only But no
sulphur Proteins contain sulfur but no
phosphorous
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2nd Proof DNA acts as a genetic material Alfred
Hershey Martha Chase (1952) Studied E.coli
cells after infection with phage T2
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Hershey Chase produced phage particles
containing radioactive DNA
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• E.Coli cells grown in ³²P containing medium
  (DNA becomes radioactive)
• Infection with non-radioactive T2 phage
• DNA labeled progeny phages

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Phage particles containing radioactive
proteins were produced
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• E.Coli cells grown in ³S-
• containing medium
• Infection with non-radioactive T2 phage
• Protein labeled progeny phages

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Genetic material in T2 phage is DNA
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Experiments of Avery, Macleod
Carty AND Hershey Chase are regarded as
classics in the demostration that genes consist
of DNA
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• DNA is the genetic material in all cellular
  organisms many viruses
• A few type of viruses have RNA as the genetic
  material

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• DNA had been proven as the genetic material by
  the Hershey-Chase experiments, but how DNA served
  as genes was not yet certain
• TO SERVE AS GENE
• DNA must carry information from parent cell to
  daughter cell
• It must contain information for replicating
  itself
• It must be chemically stable, relatively
  unchanging
• However, it must be capable of mutational change
  Without mutations there would be no process of
  evolution.

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• Many scientists were interested in deciphering
  the structure of DNA
• Among them were Francis Crick, James Watson,
  Rosalind Franklin, and Maurice Wilkens

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• Franklin took X-ray diffraction photomicrographs
  of crystalline DNA extract
• These studies of Rosalind Franklin Maurice
  Wilkens show very clear picture of X-ray
  diffraction of DNA

These lines show the helical nature of DNA
molecule
Dark structures show regularities or periodicity
in helical structure ( Recurring bases)
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Base equivalence rule given by Chargaff He
analyzed the base sequence (molar proportions of
bases) of different cells, tissues and also from
different species
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Source of DNA A G C
T 5meC Bovine thymus 28.2
21.5 21.2 27.8 1.3 Bovine
spleen 27.9 22.7 20.8
27.3 1.3 Bovine sperm 28.7 22.2
20.7 27.2 1. 3 Rat bonemarrow
28.6 21.4 20.4 28.4
1.1 Herring testes 27.9 19.5
21.5 28.2 2.8 Wheat germ
27.3 22.7 16.8 27.1
6.0 Yeast 31.3 18.7
17.1 32.9 - E.Coli
26.0 24.9 25.2 23.9
- X174 24.3 24.5
18.2 32.3 - A T are present
in equimolar amounts G C are also present in
equimolar amounts
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• Thus, Chargaff made this observation that
  AT GC
• But this quantity varies from species to species
• This equivalence is of utmost importance in
  relation to the formation of DNA helix and may be
  referred to as Chargaffs rule
• Also, Sum of Purines Sum of Pyrimidines
• Sum of amino base Sum of keto (oxo) bases
• (A C)
  (G T)

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• Major deviations from the rule
• In wheatgerm DNA G C but if 5-methyl cytosine
  is added to it, the scarcity of cytosine is
  compensated and so it could be explained
• 2. In ?X174 DNA, A T G C this is because
  ?X174 DNA is single stranded
• These rules can only be applied to double
  stranded DNA and not to RNA single stranded DNA
  

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Watson and Crick gathered all available data in
an attempt to develop a model of DNA
structure
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From the available data, Watson Crick proposed
a secondary structure of DNA
Main Features of their model are
DNA molecule is double stranded and in the form
of a right handed helix with the two
polynucleotide chains wound round the same axis
and held together by H- bonds between the bases
The two polynucleotide are anti-parallel one
chain runs from 5 3 the other from 3
5 direction
The helix has two grooves- major groove minor
groove. These grooves are alternative
The backbone is made up of alternating
deoxyribose sugar and negatively charged
phosphate groups. The various bases are attached
to the sugar
The bases are hydrophobic are stacked inside
the double helix
The negatively charged phosphate group is
hydrophilic on the outside of the double helix,
facing the surrounding water
The bases are perpendicular to the long axis of
the double helix
Each turn of the helix has 10 base pairsThere is
a distance of 0.34 nm between each base pair.Thus
one complete turn of helix is 3.4 nm
The diameter of the helix is 2 nm
The bases have a rotation of 36 so after one
complete turn they rotate at 360
The DNA strands have complementary base pairings
i.e. A can pair with T G can pair with C
Three H- bonds can form between G C but only
two can form between A T
Other pairings of bases don't fit the double
helix structure
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P
P
S
S
T
A
P
P
S
S
C
G
P
P
S
S
G
C
P
P
S
S
A
T
P
P
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• The diameter of DNA helix is 20 A in this
  diameter a base pair made up of one purine and
  one pyrimidine can fit easily
• A base pair made up of two purines would be too
  large to fit inside the helix and a base pair
  made up of two pyrimidines would be too far apart
  to form stable hydrogen bond
• So to maintain regularity in structure only one
  purine can bind to one pyrimidine

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• DNA was proven as the hereditary material and
  Watson et al. had deciphered its structure.
• Now the question was how DNA copies its
  information and how that was expressed in the
  phenotype as cells multiply and give rise to new
  cells, the genome must be accurately duplicated
  so that information is passed to each new
  generation with minimal error.

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• Watson and Crick while describing the discovery
  of a model structure for B-DNA gave a famous
  statement
• It has not escaped our notice that the specific
  pairing we have postulated immediately suggests a
  possible copying mechanism for the genetic
  material.

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• Three possible modes of replication could be
  hypothesized based on Watson and Cricks model
  for the structure of the DNA double helix
• Semiconservative,
• Conservative
• Dispersive.

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SEMI-CONSERVATIVE REPLICATION
CONSERVATIVE REPLICATION
 DISPERSIVE REPLICATION
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• Watson and Crick proposed the hypothesis of
  semiconservative replication
• This hypothesis was proved by experiments
  carried out by Matthew Meselson and Franklin
  Stahl in 1957.

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• Meselson and Stahl grew E. coli cells for many
• generations in a medium in which the sole
  nitrogen source (NH4Cl) contained ¹5N, the
  heavy isotope of nitrogen, instead of the
  normal, more abundant light isotope,¹4N
• The DNA isolated from these cells had a density
  about 1 greater than that of normal ¹4NDNA and
  a mixture of heavy ¹5NDNA and light ¹4NDNA
  can be separated by centrifugation to equilibrium
  in a cesium chloride density gradient

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• The E. coli cells grown in the ¹5N medium were
• transferred to a fresh medium containing only the
  ¹4N isotope, where they were allowed to grow
  until the cell population had just doubled
• The DNA isolated from these first-generation
  cells formed a single band in the CsCl gradient
  at a position indicating that the double helical
  DNA molecules of the daughter cells were hybrids
  containing one new ¹4N strand and one parent ¹5N
  strand

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• The semi conservative replication hypothesis was
  further supported in the next step of the
  experiment
• Cells were again allowed to double in number in
  the ¹4N medium
• The isolated DNA product of this second cycle of
  replication exhibited two bands in the density
  gradient, one with a density equal to that of
  light DNA and the other with the density of the
  hybrid DNA observed after the first cell doubling

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THANK YOU