CHAPTER

21

MOLECULAR BASIS OF
INHERITANCE
 Nucleic acids (DNA & RNA) are the building blocks of genetic material.
DNA is the genetic material in most of the organisms.
RNA is the genetic material in some viruses. RNA mostly functions
as messengers.

THE DNA
STRUCTURE OF POLYNUCLEOTIDE CHAIN
Polynucleotides are the polymer of nucleotides. DNA &
RNA are polynucleotides. A nucleotide has 3 components:
1. A nitrogenous base.
2. A pentose sugar (ribose in RNA & deoxyribose inDNA).
3. A phosphate group.
Nitrogen bases are 2 types:
- Purines: It includes Adenine (A) and Guanine (G).
- Pyrimidines: It includes Cytosine (C), Thymine (T) &
Uracil (U). Thymine (5-methyl Uracil) present only in
DNA and Uracil only in RNA.
A nitrogenous base is linked to the pentose sugar through an
N-glycosidic linkage to form nucleoside.

Nucleosides in RNA Nucleosides in DNAt

Adenosine Deoxyadenosinet
Guanosine Deoxyguanosine
Cytidine Deoxycytidine
Uridine Deoxythymidine

Nitrogen base + sugar + phosphate group = Nucleotide

(deoxyribonucleotide).

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

In RNA, every nucleotide residue has an additional –OH
group present at 2'-position in the ribose (2’- OH).
2 nucleotides are linked through 3’-5’ phosphodiesterbond
to form dinucleotide.
When more nucleotides are linked, it formspolynucleotide.

STRUCTURE OF THE DNA

- Friedrich Meischer (1869): Identified DNA and named it
as ‘Nuclein’.
- James Watson & Francis Crick (1953) proposed double
helix model of DNA. It was based on X-ray diffraction data
produced by Maurice Wilkins & RosalindFranklin.

- DNA is made of 2 polynucleotide chains coiled in a righthanded

fashion. Its backbone is formed of sugar &
phosphates. The bases project inside.
- The 2 chains have anti-parallel polarity, i.e. one chain has
the polarity 5’-3’ and the other has3’-5’.
- The bases in 2 strands are paired through H-bondsforming
base pairs (bp).
A=T (2 hydrogen bonds) C-G (3 hydrogenbonds)
- Purine comes opposite to a pyrimidine. This generates
uniform distance between the 2 strands.
- Erwin Chargaff’s rule: In DNA, the proportion of A is
equal to T and the proportion of G is equal toC.

[A] + [G] = [T] +[C]

or [A] + [G] / [T] + [C] =1

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

 6 Ф 174 (a bacteriophage) has 5386nucleotides.
6 Bacteriophage lambda has 48502 base pairs(bp).
6 E. coli has 4.6x106bp.
6 Haploid content of human DNA is 3.3x109bp.

Length of DNA = number of base pairs X distance between

two adjacent base pairs.
Number of base pairs in human = 6.6 x 109
Hence, the length ofDNA = 6.6 x109 x 0.34x 10-9
= 2.2m
In E. coli, length ofDNA =1.36 mm (1.36 x 10-3m)
1 . 36x10—3
The number of base pairs =0.34 x 10—9
= 4 x 106bp

PACKAGING OF DNA HELIX

- In prokaryotes (E.g. E. coli), the DNA is not scattered
throughout the cell. DNA, being negatively charged, is held
with some positively charged proteins and form ‘nucleoid’.
- In eukaryotes, there is a set of positively charged, basic
proteins called histones.
- Histones are
in positively
rich charged
basic amino acid residues
lysines and arginines.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

- 8 histones form histone octamer.
- Negatively charged DNAis wrapped around histone octamer to give nucleo-
some.
- A typical nucleosome contains 200 bp.
Therefore, the total number of nucleosomes in human =
6.6x109bp = 3.3x107
200
- Nucleosomes constitutfeo rtmhe repeating unit to chromatin. Chromatin
is the thread-like stained bodies.

- Nucleosomes in chromatin = ‘beads-on-string’.

- Chromatin is packaged -chromatin fibres - coiled
and condensed at metaphase stage - chromosomes.
- Higher level packaging of chromatin requires nonhistone
chromosomal (NHC) proteins.
- Chromatins include
Euchromatin: Loosely packed and transcriptionally
active chromatin and stains light.
Heterochromatin: Densely packed and inactive region
of chromatin and stains dark.

THE SEARCH FOR GENETICMATERIAL

1. GRIFFITHˇS TRANSFORMING PRINCIPLE EXPERIMENT
Griffith used mice &Streptococcus pneumoniae.
Streptococcus pneumoniae has 2 strains-
- Smooth (S) strain (Virulent): Has polysaccharide
mucus coat. Cause pneumonia.
- Rough (R) strain (Non-virulent): No mucus coat. Does
not cause Pneumonia.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

 Experiment:
S-strain ---- Inject into mice ---- Mice die
R-strain ---- Inject into mice ---- Mice live
S-strain (Heat killed) ---- Inject into mice ---- Mice live
S-strain (Hk) + R-strain (live) ---- Inject into mice ---- Mice die

He concluded that some ‘transforming principle’, transferred

from heat-killed S-strain to R-strain. It enabled R-strain to
synthesize smooth polysaccharide coat and become virulent.
This must be due to thetransfer of genetic material.

2. BIOCHEMICAL CHARACTERIZATION OF
TRANSFORMING PRINCIPLE
- Oswald Avery, Colin MacLeod & Maclyn McCarty worked to determine
the biochemical nature of transforming principle’ in Griffith’s experiment.
- They purified biochemicals (proteins, DNA, RNA etc.) from heat killed S
cells using suitable enzymes.
- They discovered that
Digestion of protein and RNA (using Proteases and RNases) did not
affect transformation. So, the transforming substance was not a protein
or RNA.
Digestion of DNA with DNase inhibited transformation. It
means that DNA caused transformation of R cells to S
cells, i.e. DNA was the transforming principle.

3.HERSHEYCHASE EXPERIMENT
(BLENDERE XPERIMENT)
- Hershey & Chase grew some bacteriophage viruses on a medium contain-
ing radioactive phosphorus (P32) and some others on medium containing
radioactive sulphur (S35).

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

- Viruses grown in P32 got radioactive DNA because only DNA contains
phosphorus. Viruses grown in S35 got radioactive protein because protein
contains sulphur.
Bacteriophage Radioactive (32P)

- These preparations were used Radioactive (35S) labelled

protein capsule
labelled DNA

separately to infect E. coli.

- After infection, the E. coli cells 1. Infection

were gently agitated in a blender to

remove the virus particles from the 2. Blending

bacteria.
3. Centrifugation

- Then the culture was centrifuged to No Radiactive (35S)

detected in cells
Radioactive (32P)
detected in cells

separate lighter virus particles Radioactive

detected in supernatant
No Radiactivity
detected in supernatant

from heavier bacterial cells.

- Bacteria infected with viruses having radioactive DNA
were radioactive. i.e., DNA had passed from the virus to
bacteria. Bacteria infected with viruses having radioactive
proteins were not radioactive. i.e., proteins did not enter the
bacteria from the viruses. This proves that DNA is the
genetic material.

PROPERTIES OF GENETIC MATERIAL

(DNA v/s RNA)
A genetic material may have the following properties:
Ability to generate its replica (Replication).
Chemical and structural stability.
Provide the mutations that are required for evolution.
Ability to express as ‘Mendelian Characters’.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

 Reasons for stability Reasons formutability
(less reactivity) ofDNA (high reactivity) ofRNA

Double stranded Single stranded

Presence of thymine Presence of Uracil
Absence of 2’-OHin sugar Presence of 2’-OHin sugar

- RNA is unstable. So, RNAviruses (E.g. Q.Bbacteriophage,

Tobacco Mosaic Virus etc.) mutate and evolve faster.
- DNA strands are complementary. On heating, they separate.
In appropriate conditions, they come together. In Griffith’s
experiment, some properties of DNA of the heat killed
bacteria did not destroy. It indicates the stability of DNA.
- For the storage of genetic information, DNA is better due
to its stability. But for the transmission of genetic
information, RNA is better.
- RNA can directly code for the protein synthesis, hence can
easily express the characters. DNA is dependent on RNA
for protein synthesis.

RNA WORLD
- RNA was the first genetic material.
- It acts as genetic material and catalyst.
- Essential life processes (metabolism, translation, splicing
etc.) evolved around RNA.
- DNA evolved from RNA for stability.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

CENTRAL DOGMA OF MOLECULAR
BIOLOGY
It is proposed by Francis Crick. It states that the genetic
information flows from DNA - RNA - Protein.
In some viruses, flow of information is in reverse direction
(from RNA to DNA). It is called reverse transcription.

DNA REPLICATION
Replication is the copying of DNA from parental DNA.
Watson & Crick proposed Semi-conservative model of
replication. It suggests that the parental DNA strands act as
template for the synthesis of new complementary strands.
After replication, each DNA molecule would have one
parental and one new strand.
Matthew Messelson & Franklin Stahl (1958)
experimentally proved Semi-conservative model.

Messelson & Stahlˇs Experiment

- They cultured E. coli in a medium containing 15NH4Cl (15N: heavy isotope of
N).15N was incorporated into both strands of bacterial DNA and the DNA
became heavier.
- Another preparation containing N salts labeled with 14N is also made. 14N
was incorporated in both strands of DNA and became lighter.
These 2 types of DNA can be separated by centrifugation in a CsCl density
gradient.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

 - They took E. coli cells from 15N medium and transferred to
14N medium. After one generation (i.e. after 20 minutes),
they isolated and centrifuged the DNA. Its density was
intermediate (hybrid) between 15N DNA and 14N DNA.
This shows that the newly formed DNA one strand is old
(15N type) and one strand is new (14N type). This confirms
semi-conservative replication.
After II generation (i.e. after 40 minutes), there was equal
amounts of hybrid DNA and light DNA.

Taylor & colleagues (1958) performed similar experiments on

Vicia faba (faba beans) using radioactive thymidine to
detect distribution of newly synthesized DNA in the
chromosomes. It proved that the DNA in chromosomes also
replicate semi-conservatively.

The Machinery and Enzymes for Replication

DNA replication starts at a point called origin(ori).
A unit of replication with one origin is called a replicon.
During replication, the 2 strands unwind and separate by
breaking H-bonds in presence of an enzyme, Helicase.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

Unwinding of theDNA molecule at a point forms a structure
‘Y’-shaped called replication fork. The separated strands act
as templates for the synthesis of new strands. DNA replicates in the
5’-3’ direction.
Deoxyribonucleoside triphosphates (dATP, dGTP, dCTP
& dTTP) act as substrate and provide energy for
polymerization.

Firstly, a small RNA primer is synthesized in presence of

an enzyme, primase.
In the presence of an enzyme, DNA dependent DNA polymerase, many
nucleotides join with one another to primer strand and form a polynu-
cleotide chain (new strand).
The DNA polymerase forms one new strand (leading strand) in a contin-
uous stretch in the 5’-3’ direction (Continuous synthesis).

The other new strand is formed in small stretches (Okazaki

fragments) in 5’- 3’ direction (Discontinuous synthesis).
The Okazaki fragments are then joined together to form a
new strand by an enzyme, DNA ligase. This new strand is
called laggingstrand.
If a wrong base is introduced in the new strand, DNA
polymerase can do proof reading.
E. coli completes replication within 38 minutes. i.e. 2000
bp per second.
In eukaryotes, the replication of DNA takes place at Sphase
of the cell cycle. Failure in cell division afterDNA
replication results in polyploidy.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

TRANSCRIPTION
- It is the process of copying genetic information from one
strand of the DNA into RNA.
- Here, adenine pairs with uracil instead of thymine.
- During transcription, both strands are not copied because
- The code for proteins is different in both strands. This
complicates the translation.
- If 2 RNA molecules are produced simultaneously, this
would be complimentary to each other. It forms a double
stranded RNA and prevents translation.

TRANSCRIPTION UNIT
- It is the segment of DNA between the sites of initiation and termi-
nation of transcription. It consists of 3 regions:
- A promoter (Transcription start site): Binding site for RNA poly-
merase.
- Structural gene: The region between promoter and terminator where
transcription takes place.
- A terminator: The site where transcriptionstops.
- The DNA- dependent RNA polymerase catalyzes the
polymerization only in5’-3’direction.
- 3’-5’ acts as template strand. 5’-3’ acts as coding strand.
3’-ATGCATGCAT GCATGCATGCATGC-5’te mplate strand.
5’-TACGTACGTACGTACGTA CGTACG-3’ codingstrand.

TRANSCRIPTION UNIT ANDGENE

Gene: Functional unit of inheritance. It is the DNA sequence
coding for RNA molecule.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

Cistron: A segment of DNA coding for a polypeptide. Structural gene in a
transcription unit is 2 types:
- Monocistronic structural genes (split genes): It is seen in eukaryotes.
Here, coding sequences (exons or expressed sequences) are interruptedby
introns(intervening sequences).
- Polycistronic structural genes: It is seen in prokaryotes. Here, there are
no splitgenes

STEPS OF TRANSCRIPTION IN PROKARYOTES

- Initiation: Here, the enzyme RNA polym erase binds at the
promoter site of DNA. This causes the local unwinding of
the DNA double helix. An initiation factor(σ factor) present
in RNA polymerase initiates the RNA synthesis.
- Elongation: RNA chain is synthesized in 5’-3’ direction. In
this process, activated ribonucleoside triphosphates (ATP,
It is the sequence of nucleotides (nitrogen bases) in
mRNA
that contains information for protein synthesis (translation).
- The sequence of 3 bases determining a single amino acid
is called codon
20 types of amino acids involved in translation
1. Alanine (Ala) 11. Leucine (Leu)
2. Arginine (Arg) 12. Lysine (Lys)
3. Asparagine (Asn) 13. Methionine (Met)
4. Aspartic acid (Asp) 14. Phenyl alanine (Phe)
5. Cystein (Cys) 15. Proline (Pro)
6. Glutamine (Gln) 16. Serine (Ser)
7. Glutamic acid (Glu) 17. Threonine (Thr)
8. Glycine (Gly) 18. Tryptophan (Trp)
9. Histidine (His) 19. Tyrosine (Tyr)
10. Isoleucine (Ile) 20. Valine (Val)

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

- George Gamow suggested that for coding 20 amino
acids, the code should be made up of 3 nucleotides.
- Har Gobind Khorana devineloped the chemical method synthesizing
RNA molecules with defined combinations of
bases (homopolymers & copolymers).
- Marshall Nirenberg developed cell-free system for protein
synthesis.
- Severo Ochoa (polynucleotide phosphorylase) enzyme is
used to polymerize RNA with defined sequences in a
template independent manner.

SALIENT FEATURES OF GENETIC CODE

Triplet code (three-letter code).
61 codons code for amino acids. 3 codons (UAA, UAG&
UGA) do not code for any amino acids. They act as stop
codons (Termination codons or non-sensecodons).
Genetic code is universal. E.g. From bacteria to human
UUU codes for Phenylalanine. Some exceptions are found
in mitochondrial codons, and in some protozoans.
No punctuations b/w adjacent codons (comma less code).
The codon is read in mRNA in a contiguous fashion.
Genetic code isnon-overlapping.
A single amino acid is represented by many codons (except
AUG for methionine & UGG for tryptophan). Such codons
are called degenerate codons. Genetic code is unambiguous and specific.
i.e. one codon specifies only one amino acid.
AUGhas dual functions. It codes for Methionine and acts
as initiator codon. In eukaryotes, methionine is the first
amino acid and formyl methionine in prokaryotes.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

MUTATIONS AND GENETICCODE
- Relationship between genes & DNA are best understood by mutation
studies. Deletions & rearrangements in a DNA may cause loss or gain of
a gene and so a function.
- Insertion or deletion of one or two bases changes the reading frame
from the point of insertion ordeletion.
- Insertion/ deletion of three or its multiple bases insert or
delete one or multiple codon. Hence one or multiple amino
acids are inserted /deleted. The reading frame remains
unaltered from that point onwards. Such mutations are
known as frame-shift insertion or deletionmutations.
- It proves that codon is a triplet and is read contiguously.

TYPES OF RNA
mRNA (messenger RNA): Provide template for transla-
tion
(protein synthesis).
rRNA (ribosomal RNA): Structural & catalytic role during
translation. E.g. 23S rRNA in bacteria acts asribozyme.
tRNA (transfer RNA or sRNA or soluble RNA): Brings
amino acids for protein synthesis and reads the genetic
code.
tRNA is called adaptor molecule because ithas
An Anticodon (NODOC) loop that has bases
complementary to the codon.
An amino acid acceptor end to which amino acid binds.
Ribosome binding loop.
Enzyme binding loop.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

- For initiation, there is another tRNA called initiator tRNA.
- There are no tRNAs for stopcodons.
- Secondary (2-D) structure of tRNA looks like a cloverleaf.
3-D structure looks like inverted ‘L’.

TRANSLATION (PROTEIN SYNTHESIS)

It takes place in ribosomes. It includes 4steps:
1. Charging oft RNA 2. Initiation
3. Elongation 4. Termination

1. CHARGING (AMINOACYLATION) OFTRNA

Formation of peptide bond needs energyobtained fromATP.
For this, amino acids are activated (amino acid + ATP) and
linked to their cognate tRNA in presence of am inoacyl tRNA
synthetase. Thus, the tRNAbecomes charged.

2.INITIATION
It begins at the 5’-end of mRNA in the presence of an initiation factor.
The mRNA binds to the small subunit of ribosome. Now
the large subunit binds to the small subunit to complete the
initiation complex.
Large subunit has 2 binding sites for tRNA- aminoacyl
tRNA binding site (A site) and peptidyl site (P site).
Initiation codon for methionine is AUG. So methionyl
tRNA complex would have UAC at the Anticodonsite.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

3.ELONGATION
At the P site, the first codon of mRNA binds with anticodon
of methionyl tRNA complex.
Another aminoacyl tRNA complex with an appropriate
amino acid enters the ribosome and attaches to A site. Its
anticodon binds to the second codon on the mRNA and a
peptide bond is formed between first and second amino
acids in presence of an enzyme, peptidyl transferase.
First amino acid and its tRNA are broken. This tRNA is
removed from P site and second tRNAat the A site is pulled
to P site along with mRNA. This is called translocation.
Then 3rdcodon comes into A site and a suitable tRNA with
3rdamino acid binds at the A site. This process is repeated.

4.TERMINATION
When aminoacyl tRNA reaches the termination codon like
UAA, UAG & UGA, the termination of translation occurs.
The polypeptide and tRNA are released from the ribosomes.
The ribosome dissociates into large and small subunits at
the end of protein synthesis.
A group of ribosomes associated with a single mRNA for
translation is called a polyribosome (polysomes).
An mRNA has additional sequences that are not translated
(untranslated regions or UTR). UTRs are present at both 5’-
end (before start codon) and 3’-end (after stop codon). They are
required for efficient translation process.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

REGULATION OF GENE EXPRESSION
In eukaryotes, gene expression occurs by following levels:
1. Transcriptional level (formation of primary transcript).
2. Processing level (splicing etc.).
3. Transport of mRNA from nucleus to thecytoplasm.
4. Translational level (formation of a polypeptide).
The metabolic, physiological and environmental conditions
regulate expression of genes. E.g.
In E. coli, the beta-galactosidase enzyme hydrolyses
lactose into galactose & glucose. In the absence of lactose,
the synthesis of beta-galactosidase stops.
The development and differentiation of embryo into adult
are a result of the expression of several set of genes.
If a substrate is added to growth medium of bacteria, a set of
genes is switched on to metabolize it. It is called induction.
When a metabolite (product) is added, the genes to produce it
are turned off. This is called repression.

OPERON CONCEPT
- Each metabolic reaction is controlled by a set of
genes”
- All the genes regulating a metabolic reaction constitute an
Operon. E.g. lac operon, trp operon, ara operon,
his operon, val operon etc.
Lac Operon in E. coli: The operon controlling lactose

a) A regulatory or inhibitor (i) gene: Codes for the repressor

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

 b) 3 structural genes:
i. z gene: Codes for galactosidase (hydrolyze lactose
to galactose and glucose).
ii. y gene: Codes for permease (increase permeability of
the cell to lactose).
iii. a gene: Codes for a transacetylase.

- The genes present in the operon function together in the

same or related metabolic pathway. There is an operator
region for each operon.
- If there is no lactose (inducer), lac operon remains
switched off. The regulator gene synthesizes mRNA to
produce the repressor protein. This protein binds to the
operator genes and blocks RNA polymerase movement. So
the structural genes arenot expressed.
- If lactose is provided in the growth medium, the lactose is
transported into the E. coli cells by the action of permease.
Lactose (inducer) binds with repressor protein. So repressor
protein cannot bind to operator gene. The operator gene
becomes free and induces the RNA polymerase to bind with
promoter gene. Then transcriptionstarts.
- Regulation of lac operon by repressor is called negative
regulation.

HUMAN GENOME PROJECT(HGP)

The entire DNA in the haploid set of chromosomes of an
organism is called a Genome.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

 In Human genome, DNA is packed in 23chromosomes.
Human Genome Project (1990-2003) is the first mega
project in identifying the sequence of nucleotides and
mapping of all the genes in human genome.
Human genome contains about 3x109bp.

GOALS OF HGP
a. Identify all the estimated genes in human DNA.
b. Determine the sequences of the 3 billion chemical base
pairs that make up humanDNA.
c. Store this information in databases.
d. Improve tools for data analysis.
e. Transfer related technologies to other sectors.
f. Address the ethical, legal and social issues (ELSI) that
may arise from the project.

Methodologies of HGP: 2 major approaches.

Expressed Sequence Tags (ESTs): Focused on identifying
all the genes that are expressed as RNA.
Sequence annotation: Sequencing whole set of genome
containing all the coding & non-coding sequence and later
assigning different regions in the sequence with functions.
Procedure:
Isolate total DNA from a cell - Convert into random
fragments - Clone in suitable host (e.g. BAC & YAC) for
amplification - Fragments are sequenced using Automated
DNA sequencers (using Frederick Sanger method) -- Sequences are
arranged based on overlapping regions
Alignment of sequences using computer programs

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

 BAC= Bacterial Artificial Chromosomes
YAC= Yeast Artificial Chromosomes

HGP was closely associated with Bioinformatics.

Bioinformatics: Application of computer science and
information technology to the field of biology & medicine.

SALIENT FEATURES OF HUMANGENOME

a. Human genome contains 3164.7 million nucleotidebases.
b. Total number of genes= about 30,000.
c. Average gene consists of 3000 bases, but sizes vary.
Largest known human gene (dystrophin on Xchromosome)
contains 2.4 million bases.

d. 99.9% nucleotide bases are same in all people. Only 0.1%

(3x106 bp) difference makes every individual unique.
e. Functions of over 50% of discovered genes are unknown.
f. Chromosome I has most genes (2968) and Y has the
fewest (231).
g. Less than 2% of the genome codes for proteins.
h. Very large portion of human genome is made of Repeated
(repetitive) sequences. These are stretches of DNA
sequences that are repeated many times. They have no
direct coding functions. They shed light on chromosome
structure, dynamics and evolution.
i. About 1.4 million locations have single-base DNA
differences. They are called SNPs (Single nucleotide
polymorphism or ‘snips’).

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

DNA FINGERPRINTING
(DNAPROFILING)
It is the technique to identify the similarities and differences
of the DNA fragments of 2 individuals.
Developed by Alec Jeffreys (1985).

BASIS OF DNA FINGERPRINTING

DNA carries some non-coding repetitive sequences called
variable number tandem repeats (VNTR).
Number of repeats is specific from person to person.
The size of VNTR varies from 0.1 to 20 kb.
Repetitive DNA are separated from bulk genomic DNA as
different peaks during density gradient centrifugation.
The bulk DNA forms a major peak and the small peaks
are called satelliteDNA. Satellite DNA is classified as micro-satellites,
minisatellites etc. based on base composition (A:T rich or G:C
rich), length of segment and number of repetitive units.
VNTR belongs to mini-satellite DNA.
Any difference in the nucleotide sequence (inheritab le
mutation) observed in a population is called DNA
polymorphism (variation at genetic level).
Polymorphism is higher in non-coding DNA sequence
because mutations in these sequences may not have any
immediate effect in an individual’s reproductive ability .
These mutations accumulate generation after generation
and cause polymorphism. For evolution & speciation,
polymorphisms play important role.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001

 STEPS OF DNA FINGERPRINTING
(SOUTHERN BLOTTINGTECHNIQUE)
a. Isolation of DNA.
b. Digestion of DNA by restriction endonucleases.
c. Separation of DNA fragments by gel electrophoresis.
d. Transferring (blotting) DNA fragments to synthetic
membranes such as nitrocellulose or nylon.
e. Hybridization using radioactive labelled VNTR probe.
f. Detection of hybridized DNA by autoradiography.
The image (in the form of dark& light bands) obtained is
called DNA fingerprint. It differs from individual to
individual except inmonozygotic (identical) twins.
The sensitivity of the technique has been increased by use of
polymerase chain reaction (PCR). Therefore, DNA from a
single cell is enough for DNA fingerprinting.

APPLICATION OF DNAFINGERPRINTING
Forensic tool to solve paternity, rape, murder etc.
For the diagnosis of genetic diseases.
To determine phylogenetic status of animals.
To determine population and genetic diversities.

anand_mani16 DR. Anand Mani https://www.anandmani.com/ https://discord.io/anandmani t.me/anandmani001