Gene:

Fine Structure of
Gene
An imaginary
overview
All information of our life
is written in two Books
Two set (23 Pairs) of
Chromosomes
One of these Books of life
is written by Father
Set of chromosomes (23)
inherited from Father
The another Book is
written by Mother
Set of chromosomes (23)
inherited from Mother
 Both of these Books are
preserved in a Bookshelf
Both set of chromosomes are
preserved in a Nucleus
 Each Book of Life has 23
Chapters with same title
except chapter number 23
Ch. 1: Ch. ---:
Ch. 1: Ch. ---:
Chromosome 1 Chromosome ---
Chromosome 1 Chromosome ---
Ch. 2: Ch. 23:
Chromosome 2 Ch. 23: Ch. 2: Chromosome X
Chromosome X / Y Chromosome 2
Each Chapter (Chromosome)
has many subtitle (Gene)
Ch. 2:
Ch. 1: Chromosome 2
Chromosome 1
Gene ETM2
Gene GBA
Gene MSH2
Gene HPC1
There are two copies (allele) of each
subtitle (Gene) in a cell as each cell
contains two books of life
Ch. 2: Ch. 2:
Ch. 1: Chromosome 2 Ch. 1:
Chromosome 1 Chromosome 2
Chromosome 1
Gene ETM2 Gene ETM2
Gene GBA Gene gba
Gene msh2 Gene MSH2
Gene HPC1 Gene HPC1
Each subtitle (Gene) is written with
a 4 letters (A, T, G, C) language
Depending on external and/or internal
need, specific subtitle (Gene) is selected
for reading by reader (Cell)
Ch. 2:
Ch. 1: Chromosome 2
Chromosome 1
Gene ETM2
Gene GBA
Gene MSH2
Gene HPC1
Depending on comparative expression
power, one of the copies (allele) of specific
subtitle (Gene) become easily accessible
for reading by reader (Cell)
Ch. 2: Ch. 2:
Ch. 1: Chromosome 2 Ch. 1:
Chromosome 1 Chromosome 2
Chromosome 1
Gene ETM2 Gene ETM2
Gene GBA Gene gby
Gene MSH2 Gene MSH2
Gene HPC1 Gene HPC1
 EVOLUTION OF GENE CONCEPT
YEAR SCIENTIST GENE CONCEPT
1866 G.J. MENDEL A unit factor that
controls specific
phenotypic trait
1902 SIR A.E.GARROD One gene –one
metabolic block theory
1940 BEADLE & TATUM One gene-one enzyme
theory
 EVOLUTION OF GENE CONCEPT
YEAR SCIENTIST GENE CONCEPT
1957 U.M.INGRAM One gene-one
polypeptide theory
1960s C.YANOFSKY & Gene is a unit of
CO-WORKERS
recombination
CLASSICAL DEFINITION OF GENE
Gene is the unit of-
 Function (one gene specifies one
character),
 Recombination, and
 Mutation.
MORDERN DEFINITION OF GENE
 Unit of Genetic Information ( Unit
of DNA that specifies one
polypeptide)
 Includes coding as well as non-
coding regulatory sequences.
 Exons and Introns
 Exons are segments of a gene that encode
mature mRNA for a specific polypeptide
chain.
 Introns are segments of a gene that do not
encode mature mRNA. Introns are found in
most genes in eukaryotes and in some gene
of bacteriophage and archae .
 An eukaryotic Gene
Introns are removed from pre-mRNA to generate mRNA

exon1 intron1 exon2 intro n2 exon3

Gene:
duplex DNA
transcription

Primary transcript:
single stranded RNA
5' and 3' end processing

Precursor to cap AA AA
mRNA

splicing

mRNA cap AA AA

translation

Protein
 Types of exons
Transcription start polyA Stop
5’ GT AG GT AG GT AG GT AG
Gene 3’
promoter
Open reading frame

Initial exon
Internal exon
Terminal exon Translation Translation
Start Stop
mRNA 5’ 3’
5’ untranslated Protein 3’ untranslated
region coding region
region
lac Operon
 Operon(Gene cluster under control
of single promoter)
Structural gene- gene that codes for a
polypeptide
Promoter site- region where RNA polymerase
bind to initiate transcription of the structural
genes (STG).
Operator Site - region where the repressor
attaches to control the access to STG
Regulatory gene- codes for repressor proteins
 Bacterial Promoter

-35 box -10 or Pribnow or TATA box

ESSENTIAL FEATURES OF GENE
Determines the physical as well as
physiological characters.
Situated in the chromosome.
Occupies a specific position known
as Locus.
ESSENTIAL FEATURES OF GENE
Arranged in single linear order.
Occur in functional states called
Alleles.
Some have more than 2 alleles
known as Multiple Alleles.
ESSENTIAL FEATURES OF GENE
 Some may undergo sudden and
permanent change in expression called
as Mutant Gene (Mutation).
 May be transferred to its homologous
(Cross-over) or non-homologous
counterpart (Translocation).
 ESSENTIAL FEATURES OF GENE
 Can duplicate themselves very
accurately (Replication).
 Synthesizes a particular Protein.
 Determines the sequence of amino
acid in the polypeptide chain
ESSENTIAL FEATURES OF GENE

Average size of Prokaryotic gene is 1

kbp and have little diversity
Average size of Eukaryotic gene is 16
kbp and have great diversity
SOME TERMS RELATED TO GENE
 RECON - It is the smallest unit of
DNA capable of undergoing Crossing
Over & Recombination.
 MUTON - It is the smallest unit of
DNA which can undergo Mutation.
SOME TERMS RELATED TO GENE
 COMPLON - It is the unit of
complementation.
 CISTRON - The portion of DNA
specifying a single polypeptide
chain is termed as cistron.
Gene Cistron Relationship
 Prokaryotes : Genes and
Cistrons are equivalent
 Eukaryotes : Cistron is
equivalent to the exons
 Genetic Recombination
Genetic recombination involves the exchange of genetic material
(DNA):
- between multiple chromosomes
- between different regions of the same chromosome.
This process is generally mediated by:
- homology (homologous regions of chromosomes line up in
preparation for exchange)
- some degree of sequence identity.
However, various cases of nonhomologous recombination do exist
 Lederberg-Tatum Experiment for Genetic Recombination

Strain A: Strain B:
Grow if minimal medium Grow if minimal medium
supplemented with supplemented with threonine,
methionine and biotin. leucine and thiamine.
Davis’s U-tube experiment
 Ways of Genetic Recombination: Conjugation

Conjugation: The direct transfer of DNA (usually plasmid)

from one bacterial cell to another bacterial cell. It require
formation of a conjugation bridge between two bacterial cells
 Ways of Genetic Recombination: Transformation
Transformation: The genetic alteration of a cell resulting
from the direct uptake and incorporation of exogenous
genetic material (exogenous DNA) from its surroundings
through the cell membrane(s).
Ways of Transformation
 Ways of Genetic Recombination: Transduction

Transduction
is the process
by which
genetic
material,
e.g. DNA or
siRNA,
is
inserted into
a cell by a
virus.
 Ways of Genetic
Recombination:
independent assortment
Ways of Genetic
Recombination:
crossing-over
 Complementation test
 Occasionally, multiple mutations of a single wild type
phenotype are observed.
 The appropriate genetic question to ask is:
 whether any of the mutations are in a single gene, or
 whether each mutations represents one of the several
genes (complementation group) necessary for a
phenotype to be expressed.
 The simplest test to distinguish between the two
possibilities is the complementation test.
 Complementation test
 In complementation test, two mutants are crossed, and the
F1 is analyzed.
 If two mutants are crossed and F1 express wild type
phenotype, the phenomenon by which F1 do this is known
as Genetic complementation. It indicate that each mutation
is in one of two possible genes necessary for the wild type
phenotype.
 Alternatively, if the F1 does not express the wild type
phenotype, but rather a mutant phenotype, we conclude
that both mutations occur in the same gene.
Complementation test
 Cis and Trans position
Cis position: Genes in the cis position are on
the same chromosome of a pair of homologous
chromosomes.
Trans position: Genes in the trans position are
on the different chromosomes of a pair of
homologous chromosomes.
Wild type Mutant type

Wild type Wild type

 T4 rII system
The T4 rII system is an experimental system
developed in the 1950s by Seymour Benzer
It was developed for studying the substructure of the
gene.
This experimental system is based on genetic crosses
of different mutant strains of bacteriophage T4,
Bacteriophage T4 is a virus that infects the bacteria E.
coli.
 Transposons (Jumping Genes)
Transposons or Jumping genes or Movable genes can be
defined as small, mobile DNA sequences that:
 move around chromosomes with no regard for
homology and
 insertion of these elements may produce deletions,
inversions, chromosomal fusions and even more
complicated rearrangements
 Characteristics of Transposable Elements
1. They are found to be DNA sequences that code for enzymes
which bring about the insertion of an identical copy of
themselves into a new DNA site
2. Transposition events involves both recombination and
replication process which frequently generates two daughter
copies of the original transposable elements. One copy remains
at the parent site while the other appears at the target site (on
the host chromosome)
Characteristics of Transposable Elements (cont.)
3. The insertion of transposable elements invariably disrupts the
integrity of their target genes.
4. Since transposable elements carry signals for the initiation of
RNA synthesis, they sometimes activate previously dormant genes.
5. A transposable elements is not a replicon, thus, it can not
replicate apart from the host chromosome, the way that plasmid
and phage can.
6. No homology exists between the transposons and the target site
for its insertion. Many transposons can insert at virtually any
position in the host chromosome or into a plasmid.
Types of Transposable elements
Transposable elements can be classified into several
types, but broadly two types:
1. Insertion sequence or simple transposons
2. Composite or complex transposons
 Insertion sequence or simple transposons
 An insertion sequence is a short DNA sequence that acts as a simple transposable element.
 Insertion sequences have two major characteristics:
 they are small relative to other transposable elements (generally around 700 to 2500 bp in
length) and
 only code for proteins implicated in the transposition activity
 These proteins are usually the transposase which catalyses the enzymatic reaction allowing
the IS to move, and also one regulatory protein which either stimulates or inhibits the
transposition activity.
 The coding region in an insertion sequence is usually flanked by inverted repeats.
 In addition to occurring autonomously, insertion sequences may also occur as parts of
composite transposons; in a composite transposon, two insertion sequences flank one or
more accessory genes, such as an antibiotic resistance gene (e.g. Tn10, Tn5).
Insertion sequence or simple transposons
 Composite or complex transposons
 Composite transposons (complex transposons) include extra genes sandwiched between
two insertion sequences.
 Composite transposons may help bacteria adapt to new environments.
 Repeated movements of resistance genes by composite transposition may concentrate several
genes for antibiotic resistance onto a single R plasmid.
 Nevertheless, there exist another sort of transposons, called unit transposons, that do not
carry insertion sequences at their extremities (e.g. Tn7).
 Genetic Code
 The genetic code is a set of rules defining how the four-letter (A,
T, G, C) code of DNA is translated into the 20-letter code of
amino acids, which are the building blocks of proteins.
 The genetic code is a collection of three-letter combinations of
nucleotides called codons, each of which corresponds to a
specific amino acid or to translational signal.
 Genetic Code
 The concept of codons was first described by Francis Crick and his
colleagues in 1961.
 Any altered codon (triplet of DNA nucleotides) that encodes an
incorrect amino acid or stop signal, resulting in an altered or non-
functioning peptide or protein product is known as missense
codon.
 Basis for Cryptoanalys
 Cryptoanalys is the analysis a secrete code language.
 Genetic information is written in DNA.
 DNA molecule consists of:
 Deoxyribose sugar
(One type; Arrangement diversity not possible)
 Phophate
(One type; Arrangement diversity not possible)
 Nitrogenous bases
(Four types: A, T, G, C; Arrangement diversity possible)
 Size of Codon
How 4 letters-language of DNA is translated into 20-letters language
of protein?
Explained by George Gamov (1954) by logical reasoning
 Singlet codon ?
(Maximum 4 types of codon for amino acids; Not
sufficient for 20 amino acids)
 Doublet codon ?
(Maximum 16 types of codon for amino acids; Not
sufficient for 20 amino acids)
 Triplet codon ?
(Maximum 64 types of codon for amino acids; Sufficient for
20 amino acids)
 Size of Codon
How 4 letters-language of DNA is translated into
20-letters language of protein?
 Singlet codon ?
(Maximum 4 types of codon for amino acids;
Not sufficient for 20 amino acids)
 Doublet codon ?
(Maximum 16 types of codon for amino acids;
Not sufficient for 20 amino acids)
 Triplet codon ?
(Maximum 64 types of codon for amino acids;
Sufficient for 20 amino acids)
Genetic code
 Characters of genetic code
 The code is triplet: Each codon consists of three bases (triplet). There are 64
codons. 61 codons code for amino acids.
 There is one start codon (initiation codon): AUG acts as start codon. AUG
code for methionine. Protein synthesis begins with methionine (Met) in
eukaryotes, and formylmethionine (fmet) in prokaryotes.
 Some codons acts as stop codons: These three (UAA, UGA, UAG) are stop
codons (or nonsense codons) that terminate translation.
 The code is unambiguous: Each codon specifies no more than one amino
acid.
 The code has polarity: They are all written in the 5' to 3' direction.
 Characters of genetic code
 The code is degenerate: More than one codon can specify a single amino
acid.
 All amino acids, except Met and tryptophan (Trp), have more than one
codon.
 For those amino acids having more than one codon, the first two bases
in the codon are usually the same. The base in the third position often
varies (Wobble hypothesis).
 The code is almost universal: (the same in all organisms). Some minor
exceptions to this occur in mitochondria and some organisms.
 The code is commaless (contiguous): There are no spacers or "commas"
between codons on an mRNA.
 The code is non-overlapping: Neighboring codons on a message are non-
overlapping.
Decoding genetic code by using
mini-messenger in filter binding
 Exception of Universality of Code
Codon Mammalian Yeast Universal
Mitochondria Mitochondria Code
Code Code
UGA Tryptophan Tryptophan Stop
AUA Methionine Methionine Isoleucine
CUA Leucine Threonine Leucine
AGA Stop Arginine Arginine
AGG
Differences between “Codon” and “Anticodon”
Codon:
1. It is found in DNA and mRNA.
2. Codon is complementary to a triplet of
template strand.
3. It determines the position of an amino acid
in a polypeptide.
Anticodon
1. It occurs in tRNA.
2. It is complementary to a codon.
3. It helps in bringing a particular amino acid
at its proper position during translation.
Wobble hypothesis
 Regulation of Gene Action
 The synthesis of particular gene products is controlled by mechanisms
collectively called regulation of gene action.
 Synthesis of gene products can be controlled at the level of-
- Genome (DNA) (usually in eukaryotes)
- Transcription
- Post-transcription (usually in eukaryotes)
- Translation
- Post-translation
 Regulation of Gene Action at the Level of Genome
At the level of genome, the following five modes of regulation are operative:
1. Situation of total genetic shutdown. Example:
(a) During mitotic phase of the cell cycle, chromatin is highly condensed to
form chromosome resulting in suspension of transcriptional activity of all
genes.

(b) In mammalian female, one of the

two X chromosomes present in somatic cells
undergoes condensation in early embryonic
stages to become Barr body resulting in
inactivation of all genes of that chromosome
(Dosage compensation).
 Regulation of Gene Action
at the Level of Genome
2. Evidences for constitutive expression of some genes.
Example- Housekeeping genes:
In molecular biology, housekeeping genes are typically constitutive genes that
are required for the maintenance of basic cellular function, and are expressed
in all cells of an organism under normal and patho-physiological conditions.
Example: gene for B-actin.
 Regulation of Gene Action
at the Level of Genome
3. Many genes are expressed only in certain tissue.
Example- Smart genes or Luxury genes:
These genes are tissue-specific or organ-specific, which means they are not
expressed in all cells. They are expressed only in certain type of cell or tissue.
They are not constantly expressed, they express only when their function is
needed. Examples of luxury genes are genes coding for heat-shock proteins.
 Regulation of Gene Action at the Level of Genome
4. Some DNA is never transcribed
in any cell.
Example- Centromere
of chromosome

5. Some DNA is spliced to cause

gene rearrangement.
Example- Such a mechanism
occurs during expression of
immunoglobulin (Ig) genes.
Regulation of Gene Action
at the Level of Transcription
 Autoregulation

mRNA mRNA

Autoregulation of gene action occurs, when the product of a gene activates

or repress its own production. Two types: Positive autoregulation (the
product of a gene activates its own production) and Negative
autoregulation (the product of a gene represses its own production)
Positive and Negative Regulation of gene expression
 Negative Regulation:
Inducible System (Lac Operon)
BPs +/- 111 - 35 -26 0 3063 800 800
Regulator Promoter Operator Lac Z Lac Y Lac A
Peptide
Amino acid 360 1021 275 275
MW (Da) 3800 1,25,000 30,000 30,000

Active
Protein Tetramer Tetramer Monomer Dimer

Function Repressor β- Galactosidase β- Galactoside β- Galactoside

Permease Trans acetylase
Regulatory gene:
 Negative Regulation: Repressible System
A repressible system in Salmonella typhimurium
Regulatory gene

Utilized by (Excess)
the cell Histidine Corepressor Aporepressor

Repressor
Enzymes Genes
Metabolites

e10 g10
10
e9 g9

9
..
. e1 to e8 g1 to g8
1
 Positive Regulation: (Lac Operon)
BPs +/- 111 - 35 -26 0 3063 800 800
Regulator Promoter Operator Lac Z Lac Y Lac A
RNA-
Pol Rp
cAMP CAP β- Galactosidase β- Galactoside β- Galactoside
RNA-
Pol
Permease Trans acetylase
Rp

Lac Lac
cAMP CAP
Lac

CAP
cAMP Glucose + Galactose
Indirectly inhibit synthesis

CAP= Catabolic activator Protein

cAMP= Cyclic Adenosine Mono Phosphate
Britten-Davidson model
Regulation of Gene Action at Post-transcription level (in eukaryotes)
Expression of a gene can be regulated in
post-transcription level in following
ways:
1. By controlling mRNA processing
mechanisms such as Capping,
Splicing and 3’-polyadenylation. Only
25% of pre-mRNA can be selected for
processing.
2. By controlling the mRNA export from
nucleus.
3. By RNA editing
4. By modifying mRNA stability
Regulation of Gene Action at Post-transcription level (in eukaryotes)

1. a) Capping:
Capping changes
the five prime end
of the mRNA to a
three prime end by
5'-5' linkage, which
protects the mRNA
from 5' exonuclease,
which degrades
foreign RNA. The
cap also helps in
ribosomal binding.
Regulation of Gene Action at Post-transcription level (in eukaryotes)

1. b) Splicing:
Splicing removes the introns,
noncoding regions that are
transcribed into RNA, in order to
make the mRNA able to create
proteins. Cells do this by
spliceosomes (composed of small
nuclear ribonucleoproteins,
snPNPs) binding on either side of
an intron, looping the intron into
a circle and then cleaving it off.
The two ends of the exons are
then joined together.
Regulation of Gene Action at Post-transcription level (in eukaryotes)

1. c) 3’ Polyadenylation:
By Polyadenylation , a stretch
of RNA that is made solely of
adenine bases is added to the
3' end, and acts as a buffer to
the 3' exonuclease in order to
increase the half life of
mRNA.
 Regulation of Gene Action at Post-transcription level (in eukaryotes)

2. By controlling the mRNA export

from nucleus:
 After processing mRNA export
from nucleus to cytoplasm which
is mediated by certain proteins,
factors and receptors.
 The RNA export from nucleus to
cytoplasm is strictly regulated.
 Only 5% of heterogeneous
nuclear RNA (hnRNA) can be
exported from nucleus to
cytoplasm.
 Regulation of Gene Action at Post-transcription level (in eukaryotes)

3. By RNA editing : RNA editing is a

molecular process through which
some cells can make discrete changes
to specific nucleotide sequences
within a RNA molecule after it has
been generated by RNA polymerase.
RNA editing in mRNAs effectively
alters the amino acid sequence of the
encoded protein so that it differs from
that predicted by the genomic DNA
sequence. Exception: It can be found
in eukaryotes and their viruses, and
prokaryotes.
Regulation of Gene Action at Post-transcription level (in eukaryotes)

4. By modifying mRNA stability:

 mRNA Stability can be manipulated in order to
control its half-life.
 Stable mRNA can have a half life of up to a day or
more which allows for the production of more
protein products.
 Capping, the poly(A) tail has some effect on this
stability, as previously stated.
 Regulation of Gene Action at Translation level
In prokaryote:
 Life-time of mRNA is genetically predetermined. But, the life time is
correlated with number free ribosomes available at a given moment. Hence,
bacteria can modify their protein synthesis by altering their ribosomal
contents.
 Protein synthesis is determined by the location of a gene in a polycistronic
mRNA (polarity gradient). eg. lac Z, lac Y and lac A protein synthesis rate is
1 : 0.5 : 0.2 respectively.
In eukaryotes:
 Extension of life-time of mRNA: Life-time of mRNA can be increased by
masking it with protein particles. eg. Informosomes or masked mRNA.
 Regulation of rate of protein synthesis with recruitment factors which
apparently interferes with formation of the ribosomes-mRNA complex.
 Regulation of Gene Action at Post-translation level

 Some proteins are altered

after synthesis, usually by
partial degradation or
trimming, to form active
form of protein.
 For example, central section
of the proinsulin molecules
is removed by the
enzymatic action to yield
the active protein, insulin.