this file includes the packaging of DNA in Prokaryotic and Eukaryotic cell, the difference between DNA and RNA, Semi-conservative mode of DNA replication, and the mechanism of DNA replication.

1. DNA PACKAGING
The distance between two consecutive base pairs = 0.34nm = 0.34×10 -9 m
Total number of base pairs (diploid content) = 6.6 × 109 bp
The length of DNA double helix = 0.34 × 10-9 × 6.6 × 109
= 2.2 m approx.
The dimension of a typical nucleus = 10-6 m
It is difficult to accommodate such a long polymer inside the cell.
Hence, packaging is required.

2. 1. If the length of E. Coli DNA is 1.36 nm, calculate the number of base
pairs?
2. In an experiment, DNA is treated with a compound which tends to place
itself amongst the stacks of nitrogenous base pairs. Sa a result of which the
distance between two consecutive base increases, from 0.34nm to 0.44nm.
Calculate the lengthy of DNA double helix (having 2×109 base pairs) in the
presence of saturating amount of this compound.
3. Calculate the length of DNA of bacteriophage lambda. (48502) bp.

3. DNA packaging in PROKARYOTES
 Prokaryotes lack well defined nuclear membrane, but DNA is not scattered
throughout the cell.
 DNA being negatively charged is coiled around some positively charged
proteins is a specific region called nucleoid.
 The DNA in nucleoid is organised in large loops held by proteins.

4. DNA packaging in EUKARYOTES

5.  There is a set of four positively charged basic proteins called HISTONES. These are rich in
lysine and arginine,
 Four types of histones [ H2A, H2B, H3, and H4] occur in pair to form the CORE of nucleosomes.
 H1 is the linker that connects the DNA from adjacent nucleosomes.
 Histones are organised to form a unit of 8 molecules called histone octamer.
 The DNA (negatively charged) is wrapped around the histone octamer (positively charged) to
form a nucleosome. [200bp]
 Nucleosome constitute the repeating unit of a structure in nucleus called chromatin.
 Chromatin is thread-like stained coloured bodies inside the nucleus.
 Chromatin appears as a bead on a string like structure under the microscope.

6.  The chromatin is packed to form a solenoid structure of 30 nm diameter.
 The further supercoiling tends to form a looped structure called chromatid.
 This further coils and condensed at metaphase stage of cell division to form
chromosome.
 Non Histone Chromosomal proteins (NHC) – additional set of proteins that contribute
to packaging at higher level.
 Chromatins are of two types: EUCHROMATIN and HERTROCHROMATIN.

7. THE RNA
 Was the first genetic material.
 Evidences- essential life processes such as metabolism, translation, splicing etc.
evolved around RNA.
 RNA used to act as a genetic material as well as catalyst. But being catalyst, it was
reactive and hence unstable.
 Therefore, DNA has evolved from RNA with chemical modifications that make it more
stable.

8. DNA v/s RNA
DNA RNA
DNA contains A, T, G, and C nitrogenous
bases.
RNA contains A, U, G,and C nitrogenous
bases.
DNA is a double stranded molecule. RNA is the single stranded molecule.
Contains deoxyribose sugar. Contains ribose sugar.
DNA is a genetic material is highly
stable due to double-helical structure
and absence of oh group at 2’ carbon
position.
RNA is reactive due to 2’OH group
present at every nucleotide.
DNA is highly stable and hence cannot
act as a catalyst.
RNA is reactive and hence, possess
catalytic properties.
Mutates slowly Have high mutation rate.

9. DNA REPLICATION
 Watson and Crick in 1953 proposed that DNA replicates Semi-
conservatively.
 They quoted “ it has not escaped our notice that the specific pairing
we have postulated immediately suggests a possible copying
mechanism for the genetic material”.
 This scheme states:
i. The two strands would separate and act as a template for the synthesis
of new complementary strand.
ii. After replication, each molecule would have one parental and one
newly synthesised strand.
iii. This scheme was termed as SEMI CONSERVATIVE DNA replication.

10. THE EXPERIMENT

11. Experimental proof
 MATHEW MESELSON and FRANKLIN STAHL performed an experiment on E. coli to
prove that DNA replicates semi-conservatively.
i. They grew E.coli in a medium containing 15NH4Cl as the only nitrogen source
for many generations. As a result 15N was incorporated into newly synthesised
DNA. This heavy DNA was separated from normal DNA by caesium chloride
(CsCl) density gradient.
ii. Then they transferred the cells into a medium with normal 14NH4Cl as a result
14N was incorporated into the newly synthesised DNA strand. Then they took
samples at various time intervals.
iii. Various samples were separated independently on CsCl gradient to measure
the density of DNA.
iv. The DNA was extracted from the culture after one generation (20 min) and a
band of hybrid density was formed.
v. The DNA extracted from the culture after two generation (40 min) composed
of equal amount of hybrid DNA and light DNA.

12. If E.coli was allowed to grow for 80 minutes
then what would be the proportions of light and
hybrid densities DNA molecule?
TAYLER and COLLEAGUES (1958) – they performed similar experiments on Vicia
faba (faba beans) using radioactive thymidine to detect the distribution of newly
synthesised DNA in the chromosomes.

13. The ENZYMES for DNA replication
The process of replication requires a set of catalyst (enzymes)
 DNA-Dependent DNA polymerase
i. main enzyme to catalyse the polymerisation of deoxynucleotides.
ii. The average rate of polymerisation is 2000bp/sec.
iii. In prokaryotes- there are three types of DNA polymerases- I, II, and III. Polymerase III is
the main enzyme.
iv. In eukaryotes- there are five types of DNA polymerases- α, β, γ, δ, and ε. Polymerase
δ is the main enzyme.

14. These enzymes are highly efficient and have high accuracy. Any mistakes would lead to
mutations. Other than polymerization, these enzymes are involved in proof reading.
 HELICASE
i. It unwinds the DNA strand to form replication folk.
ii. It removes all the hydrogen bonds between the complementary strands.
iii. It also releases tension created by unwinding of DNA helix.
 TOPOISOMERASE
i. Also called molecular scissors.
ii. It cuts open the DNA by removing phospho-diester bonds between adjacent
nucleotides.
 SSbp (Single Stranded binding proteins)
i. It prevents the rewind of Single stranded DNA.
 DNA LIGASE
i. Also called molecular glue.
ii. Joins two strands of DNA together.
iii. Repair single strand breaks in duplex DNA.

15. THE PROCESS
1. The replication begins at ori (origin of replication).
2. The topoisomerase cuts the DNA at ori.
3. The helicase separates the helix by removing all the H-bonds. It also releases tension
created by unwinding.
4. As a result we end up with two Y shaped replication folks.

16. 5- The helices are prevented to rebind by Single-stranded binding proteins.
6- the DNA-dep. DNA-polymerase adds dNTPs deoxyribonucleotides triphosphates to
catalyse the growing polynucleotide. dNTPs acts as a template and provides energy for the
process.

17. 7- the DNA replication always proceeds in 5’ 3’ direction. So, parental strand with
polarity 3’ 5’ acts as a template.
8- RNA-primer is needed for initiation. It provides free 3’-OH group.
9- DNA polymerase add deoxyribonucleotides to free 3-end of the polynucleotide.
10- the replication of 3’ 5’ is continuous and is called leading strand.
11- the replication of 5’ 3’ is discontinuous and is called lagging strand.
12- the oka-zaki fragments are later joined by DNA ligase enzyme.