Maharashtra Board XII the Science (Biology) Chapter 4 (Part 1)

1. •
MOLECULAR BASIS OF INHERITANCE
Biology (XII)–
Chapter 4
(Part –1)
By: Dr. Janaki V.
Pandey (Ph.D.)

2. THE DISCOVERY OF DNA
• Friedrich Miescher
isolated a cellular substance
from the nuclei of pus cells
and called it Nuclein.
• Nuclein has a high
Phosphorus content so shows
acidic properties.
Hence it was named as nucleic
• There are two kinds of nucleic
acids found in cells i.e
DNA(deoxyribonucleic acid)
and RNA (ribonucleic acid)

3. GRIFFITH’S EXPERIMENT
• In 1928, Frederick Griffith
performed an experiment
on bacterium Streptococcus
pneumoniae that causes
Pneumonia
• Griffith used two strains of
Streptococcus i.e
1. Virulent, Smooth, pathogenic and
encapsulated S-type
2. Non-virulent, Rough, non-
pathogenic and non-
capsulated R-type.

4. • He collected S type bacteria from the blood of these dead mice.
• Genetic material from the heat killed S strain changed the R type bacteria
into S type bacteria.
Griffith called this as “transforming principle”.

5. AVERY, McCARTY MACLEOD’S EXPERIMENT
• Proved that DNA is the genetic material.
• They purified DNA, RNA, proteins and other from cell free extract of S strain
and mixed it with heat killed S strains and R strains separately.
• Only DNA was able to transform harmless R strain into pathogenic S-strain.

6. • When DNA was digested with enzyme Dnase transforming
principle did not take place.
These experiments prove that the transforming
principle is DNA but all biologists were not
convinced.

7. HERSHEY-CHASE EXPERIMENT
• Both worked with viruses that infect
bacteria i.e bacteriophages, which are
composed of DNA and protein.
• They used radioactive phosphorus (32-P)
in the medium for some viruses and
radioactive sulphur (35-S) for some others.
• DNA contains P but no S. So viruses grown
in radioactive P contained radioactive DNA
but has no radioactive protein.
• Protein have S but no P. So viruses grown
in radioactive S contained radioactive
proteins but no radioactive DNA.

8. • Radioactive bacteriophages were
allowed to infect normal E-coli
bacteria.
• It was found that bacteria infected
with radioactive DNA were
radioactive.
• Bacteria infected with radioactive
proteins were not radioactive.
• This proved that DNA is the genetic
material.

9. PACKAGING OF DNA IN PROKARYOTES
1. Eg. E.coli, cell size is almost 2-3µ long.
• 2. No well organized nucleus.
• 3. No nuclear membrane and nucleolus.
4. The nucleoid is small, circular, highly folded naked ring of DNA which is 1100µ
long and contains 4.6 million base pairs.
5. The –vely charged DNA assisted by RNA connectors and +vely charged histone like
proteins and enzymes (DNA Gyrase and Topoisomerase I) form loop like
structure for maintaining supercoiled state.

10. PACKAGING OF DNA IN EUKARYOTES
• The organization of DNA is
complex.
• Histones are required for the
packaging of DNA.
• Histones are proteins that are rich in
the basic amino acid residues lysine
and arginines which carry positive
charge in their side chain.
• Eight molecule of histones (two each of H2A, H2B, H3 and H4) get
organized to form histone octamer.
• DNA is negatively charged and it is wrapped around the
positively charged histone octamer forming a structure known
as Nucleosome.
• H1 protein binds the DNA thread where it enters and leaves the
Nucleosome.

11. • Under the electron microscope,
nucleus shows Chromatin
network.
• The nucleosomes in Chromatin
are seen as Beads-on- string’.
• Around the octamer, Core DNA is
wrapped as 1 and 3/4th turn.
• The length of Core DNA is 146 bp
(base pairs).
• Adjacent Nucleosomes are linked with Linker DNA; of
about 54 bp.
• This ‘beads-on-string’ structure gets condensed into
nucleosome fiber which is coiled like a telephone wire to
make Solenoid fiber with diameter 30nm or 300Å.

12. DNA
double
helix

13. • Additional set of
Non-Histone Chromosomal
(NHC) proteins are required
for packaging of chromatin
at higher levels.
• A loosely packed region of chromatin that stains
light, is called Euchromatin and densely packed
region that stains dark is called Heterochromatin.
• Euchromatin is considered as transcriptionally
active chromatin, while Heterochromatin is
inactive.

14. DNA REPLICATION
• All the activities of the cell are regulated and
controlled by the DNA molecule. It is carrier of
genetic information and performs two important
functions:-
• Heterocatalytic function:-
Synthesis of chemical molecules other than itself.
Eg. Synthesis of RNA(Transcription),
synthesis of protein (Translation), etc.
• Autocatalytic function:-
Synthesis of DNA itself. DNA duplicates itself by
process of replication.

15. • Through replication, it forms
two copies that are identical.
• Replication occurs in the S-
phase of Interphase in the
cell cycle and only once.
• It is Semiconservative mode
of Replication and proposed
by Watson and Crick.
• The process of replication is
as below:-

16. 1. Activation of Nucleotides:-
• 4 types of DNA nucleotides are present in the
Nucleoplasm- dAMP, dGMP, dCMP and dTMP.
• They are activated into triphosphates like
dATP, dGTP, dTTP and dCTP using ATP in the
presence of enzyme phosphorylase. The process is
known as Phosphorylation.

17. 2.Point of origin or Initiation point
• DNA replication begins at certain specific sites called ‘O’-Origin.
• In prokaryotes, there is only one origin however in Eukaryotes,
there are more than one origin.
• At the point ‘O, enzyme Endonuclease nicks one of the strands of
DNA, temporarily.
• The nick occurs in the sugar-phosphate back bone or the
phosphodiester bond.
Endonuclease

19. 3. Unwinding of DNA molecule
• The enzyme DNA helicase breaks the
hydrogen bonds in the vicinity
of ‘O’.
• The strands of DNA separate
and unwind.
• DNA molecule appears as
inverted ‘Y’- shaped structure
called Replication fork.
• Recoiling is prevented by SSBP
(single strand DNA binding
protein).
4. Replicating fork
• Stress imposed on Y-shaped structure is relieved by super helix relaxing
enzyme.

20. 5. Synthesis of new strands
• Each separated strand acts as a
template or mould.
• RNA Primer get attached at the 3' end of
template strand.
• It attracts complementary nucleotides
(dATP, dGTP, etc) from surrounding
Nucleoplasm.
• These nucleotides bind to the template
by H-bonds and are interconnected by
phosphodiester bond.
• New complementary strand is
catalyzed by DNA Polymerase.
• Always formed in 5‘--->3' direction.

21. 6. Leading and lagging strand
• Leading template- strand with free 3' end.
• Lagging template – strand with free 5' end.
• The process of replication always start at C-3 end of template and
proceed towards C-5 end.
• Both the strands of the parental DNA are antiparallel.
• New strands are always formed in 5 ‘---> 3' direction.
• Strand which develops continuously towards replicating fork is called
leading strand.
• The other develops discontinuously away from the replicating fork is called
Lagging strand.

22. • DNA synthesis on lagging template takes place in the form of small
fragments, called Okazaki fragments.
• Okazaki fragments are joined by enzyme DNA Ligase.
• RNA primers are removed by DNA polymerase and replaced by DNA
sequence with the help of DNA polymerase – I in Prokaryotes and DNA
polymerase – α in Eukaryotes.
• Finally, DNA Gyrase enzyme forms double helix to form daughter DNA
molecules.

23. 7. Formation of daughter DNA molecule
• At the end of the replication, two daughter
DNA molecules are formed.
• In each daughter DNA, one strand is parental
and the other one is totally newly synthesized.
Thus, 50% is contributed by mother DNA.
• Hence, it is described as Semiconservative
replication.

24. Enyzmes used in DNA Replication
1. Phosphorylase- activation of nucleotide
2. Endonuclease- cuts the DNA strand at phosphodiester bond.
3. Helicase- cuts the H bond.
4. SSB Protein- prevent recoilig of DNA strand.
5. Primase-synthesis RNA primer.
6. Super helix relaxing enzyme- reduces stress on replicating
fork.
7. DNA polymerase- catalyses the synthesis of new DNA strand.
8. DNA ligase-joins the okazaki fragments.
9. DNA polymerase I and ∝- used to replace RNA primer by
DNA sequence in prokaryotes and eukaryotes respectively.
10. DNA gyrase- forms double helix daughter molecule.
11. DNA topoisomerase – controls the topology of DNA during
replication.

25. Experimental confirmation of DNA Replication
• DNA replication is Semiconservative.
• Matthew Meselson and Franklin Stahl used equilibrium – density
– gradient – centrifugation technique.
• They cultured E.coli
bacteria 1st in the
medium containing 14N
(light nitrogen) and later
transferred in N15 (heavy
nitrogen) and allow to
replicate for several
generation.
• They obtained equilibrium
density gradient band by
using 6M CsCl2.
• The position of this bands
were recorded.
N15

26. • The heavy DNA (15N) molecule can be distinguished from normal DNA
by centrifugation.
(The density gradient value of 6M CsCl2 and 15N DNA is almost same.
Therefore, at the equilibrium point 15N DNA will form a band. )
• The E.coli grown in 15N will show both the strands of DNA labeled
with 15N.
• Such E.coli cells were then transferred to another medium
containing 14N normal (light) nitrogen.
• After 1st generation, one density gradient band for 14N 15N was
obtained and its position was recorded.
• After 2nd generation, two density gradient bands were obtained.
• One at 14N 15N position and other at 14N position.
• The position of bands after two generations clearly proved that
DNA replication is Semiconservative.