GENETICS

Friday, April 30, 2021
GENETICS
DR NILESH KATE
MBBS,MD
PROFESSOR
DEPT. OF PHYSIOLOGY

OBJECTIVES
 Structural & functional characteristics.
 Chromosomes
 Structure & function of DNA & RNA
 Genes.
 Applied genetics.
 Molecular genetics & Biotechnology.
 Molecular genetics & Medicine.

INTRODUCTION.
 Foundation for genetics
laid down by Mendel’s
work published in 1866.
 Demonstrated that
characteristics do not
blend but transfer
from parents to
offspring in pairs
containing single unit.

HISTORY
 Dannish botanist
Johannied called these
– Genes (1990)
 Morgan (1992) shown
to be present on
chromosomes.

STRUCTURAL & FUNCTIONAL
CHARACTEIRISTICS.
 CHROMOSOMES.
 Waldeyer (1888) coined
term Chromosomes for
thread like structure
present in Nucleus.
 Total number 46.(23
pairs)

MORPHOLOGY OF
CHROMOSOMES.
 Composed of 2
Chromatids connected at
centromere.
 Each Chromatids consists
of 2 chromonemes.
 Telomeres are terminal
end of chromosomes.

TYPES
 Morphological types
 Metacentric
 Sub Metacentric
 Acrocentric
 Telocentric
 Functional types.
 Autosomes
 Sex chromosomes.
 Supernumerary or
redundant.

MORPHOLOGICAL TYPES

CHEMICAL STRUCTURE OF
CHROMOSOME
 Mainly composed of
DNA & RNA, Basic
proteins Histones,
complex proteins,
organic phosphorus
compounds & inorganic
salts.

STRUCTURE OF DNA
 Deoxyribonucleic
acid.
 Reserve bank of genetic
information.
 Polymer of 4
monomeric
deoxyribonucleotides –
dAMP,
dGMP,dCMP,dTMP

STRUCTURE OF DNA
 Each
deoxyribonucleotide
composed of -A
nitrogen base – Purines
& Pyrimidine.(A,G,C &
T), pentose sugar &
phosphate.
 Chargaff’s rule – A=T,
G=C

WATSON-CRICK MODEL OF
DNA
 Double helix structure
with 2
polydeoxyribonucleotide
chains
 Anti-parallel chains.
 Dimensions – width 2
nm, each helix contains
10 pairs of nucleotide.

WATSON-CRICK MODEL OF
DNA
 Base with hydrogen
bonds and sugar
phosphate backbone.
 Conplementary chains
 Genetic Information –
resides in 1 strand –
template strand (sense)
other is antisense
strand.

SIZE OF DNA
 23 haploid chromosomes
have 2900000 kb with
total length 990nm.
 Thus cell contains 2m
DNA in 46 chromosomes.
 4.8 cm in each
chromosomes.

ORGANIZATION OF DNA IN
CELL
 Each DNA combines with
group of 8 histone –
nucleosomes.
 Nucleosomes & DNA packed
together to form – Solenoid
fiber
 The Solenoid Fiber in turn coil
to form chromatin fibre.
 Which is further coiled to form
Chromatin In which form DNA
is present in chromosome.

STRUCTURE OF RNA
 Polymer of Ribonucleotide
held together by 3’5’-
Phosphodiaster bridges.
 Single strand
 Sugar molecule – Ribose
 Base – Pyrimidine Base is
Uracil in place of
Thymidine.
 Chargaff’s rule – not
obeyed.

TYPES OF RNA
 Messenger RNA
(mRNA)
 Transfer RNA (tRNA)
 Ribosomal RNA
(rRNA)

DNA REPLICATION
 Process by which each
DNA gives rise to 2
copies of same.
 Method –
Semiconservation
Replication.

DNA REPLICATION
 Initiation of replication.
 Formation of
replication fork &
replication eye.
 Formation of RNA
primer.
 DNA synthesis along the
replication fork.

Initiation of replication.
 Site from which it starts called
– Origin of Replication.
 It mostly consists of A-T base
pairs.
 When sp protein (DNA
Protein) binds to site of
replication seperation of
double strand occurs
 Seperated strands form bubble
at the site of seperation.

Formation of replication fork
& Replication eye.
 Unwinding of Double
helix & forms Y shaped
Replication fork. ( if at
end) or Ө shaped
replication eye (if in
Middle)
 This step controled by
enzyme – Helicase &
single strand binding
protein.(SSB)

ROLES
 DNA Helicase
 Binds to both strands
of DNA at replication
fork & move along
double helix & separate
the strands – like
Zipper.
 Single strand
binding protein
(SSB)
 Bind to seperated
strands of DNA & keep
them separate so called
– Helix Destabilising
Proteins.
 Also provide template
for new DNA synthesis.

Formation of RNA primer.
 Required for synthesis
of new DNA.
 RNA primer
synthesized on DNA
template by RNA
polymerase.

DNA synthesis along the
replication fork.
 Occurs on both the
strands of Y shaped
DNA.
 2 types
 Continuous- in
leading strand
 Discontinuous – in
lagging.

replication fork.
 Continuous – DNA polymerase
III binds to single stranded
DNA & move along.
 Each time it meets new base on
DNA, free nucleotide approach
strand & with correct
complemetary base attach to
base in DNA by hydrogen bond.

replication fork.
 Enzyme continue to move one base at a time with new DNA
strand growing.

GENES
 General consideration.
 Genome
 Human genome
 Human genome project. (1990-2003)
 Functional genomics.
 Comparative genomics.
 Constitutive & Inducible genes.

GENES
 Functional unit of
DNA.
 Genome – total
genetic information in
a cell.

GENES
 Human genome – All the
genetic information present in
a single set of 23
chromosomes.
 Functional genomic –
understanding function of
genes & other parts of genome.

GENES ( Continue…)
 Human Genome Project(1990-2003) –
 Identified approx all 30000 genes in human DNA.
 Determined sequence of 3 billion chemical base pairs.
 Comparative genomics – analysis & comparision of
Genome from different species.

GENES ( Continue…)
 Constitutive genes.
 Product of these genes
required all the time
in cell.
 So expressed in all
cells at constant rate.
 Inducible Genes.
 Conc of proteins by these
genes are regulated by
various signals.
 Altered by Inducer or
Repressor.

GENE EXPRESSION: CENTRAL
DOGMA
 Expression of genetic material occurs through
production of proteins.
 Involves – Transcription (genetic information in DNA
transferred to RNA intermediate)& translation (
synthesis of new proteins with this information)

GENE EXPRESSION: CENTRAL
DOGMA
 This unidirectional flow of information – Central
Dogma of molecular biology given by F.H.C. Crick in
1958.

TRANSCRIPTION.
 Process in which RNA(m,t,& r)
is synthesized from DNA.
 Only for selected region of DNA.
 Strand of DNA that directs
synthesis of mRNA via
complementary base pairs –
template or coding or sense
strand.

TRANSCRIPTION.
 Other is Antisense
strand.
 Enzyme – RNA
polymerase.

TRANSCRIPTION.
 Promotor sites – RNA
polymerase binds to
promotor site – sequence
of codes on DNA –
Hogness Box/TATA Box.
 Other site CAAT site
 These helps RNA
polymerase to recognize
DNA sequence.

POST TRANSCRIPTIONAL
MODIFICATION
 mRNA is processed
from RNA transcript
by maturation.
 RNA editing also takes
place before
translation.

TRANSLATION
 Biosynthesis of proteins.
 Genetic code.
 Characteristics of genetic code.
 Process of protein biosynthesis.
 Activation of amino acids.
 Translation proper.
 Post-translational modification.

BIOSYNTHESIS OF PROTEINS.
 Process by which genetic message carried by
mRNA from DNA is converted to polypeptide
chain having specific sequence of amino
acids – Genetic Code.

GENETIC CODE.
 Information coded in mRNA is
decoded into polypeptide
chain
 Dr Hargobind Khurana
shared nobel prize with
Nirenberg & Holly for
discovering.

GENETIC CODE.
 Formed by 3 nucleotide base sequence
 Codons are made up of 4 nucleotide base
(A,G,C & U)

GENETIC CODE.
 64 different combinations
are formed, out of which
61 for 20 amino acids & 3
are termination codons
(UAA,UAG & UGA)
 AUG & GUG are initiating
codons.

CHARACTERISTICS OF
GENETIC CODE.
 Universality – same codon for same amino
acid
 Specificity- particular codon for amino acid
 Non-overlapping
 Degenerate – 1 amino acid coded by more
than 1 codons.

PROCESS OF PROTEIN
BIOSYNTHESIS.
 Activation of amino acids.
 Translation proper.
 Post-translational modification.

ACTIVATION OF AMINO ACIDS.
 Amino acids react with
ATP to form enzyme-
AMP-amino acid
complex.
 This react with specific
tRNA & amino acid is
transferred to 3’ end of
tRNA to form aminoacyl
tRNA.

TRANSLATION PROPER.
 Initiation – form
initiation complex
 Elongation – more
addition of amino acid to
carboxyl end elongate
polypeptide chain by
Ribosomes.
 Termination – stoppage
evolked by termination
codon.

POST-TRANSLATIONAL
MODIFICATION.
 It includes
 Proteolytic
degradation.
 Covalent Modification
(Phosphorylation,
Hydroxylation &
Glycosylation)

REGULATION OF GENE
EXPRESSION.
 Regulation of gene expression n
prokaryotes.
 Inducible operon system – is that regulated
genetic material which remains switched off
normally but becomes operational by Inducer.
 Catabolic pathway.
 Repressible operon system - is that regulated
genetic material which remains switched on
normally.
 Anabolic Pathway.

REGULATION OF GENE
EXPRESSION IN EUKARYOTES.
 Gene amplification.
 Gene Rearrangement.
 Regulation of gene expression through
transcription factors..
 Regulation of gene through mRNA.

Gene Amplification.
 Expression of gene increased several folds
 E.g development of drug resistance by
malignant cells to anticancer drugs

Gene Rearrangement.
 Responsible for generation of 10 billion
antigen specific immunoglobulins.

Regulation of Gene expression
through Transcription factors..
 Transcription factors are products of other
genes so mediate transregulation by binding
to specific DNA segments.

Regulation of Gene through
mRNA.
 Gene expression is
regulated by
regulation of
synthesis, transport,
processing and
stability of mRNA.

APPLIED GENETICS
 Molecular genetics & biotechnology.
 Molecular genetics & medicine.

MOLECULAR GENETICS &
BIOTECHNOLOGY.
 Genetic Engineering
 Polymerase chain reaction.
 Blotting techniques.
 Cloning.
 Apoptosis.

GENETIC ENGINEERING
RECOMBINANT DNA TECHNOLOGY.
 STAGES
 1. Generation of copy of
gene required.
 Joining the gene to a
vector or carrier
molecule.
 Introduction of vector
DNA into the host cell to
produce chimeric DNA.
 Cloning of chimeric DNA.

APPLICATION OF GENETIC
ENGINEERING
 Production of proteins of
hormones
 Molecular analysis of
disease
 Laboratory diagnostic
applications
 Gene therapy
 Prenatal diagnosis of
genetic diseases.
 Transgenesis.
 Application in forensic
medicine
 Industrial applications
 Agricultural
applications
 Evolution.

POLYMERASE CHAIN
REACTION.
 Polymerase chain
reaction is a sensitive,
selective & extremely
rapid method of
amplifying a target
sequence of DNA.
 Technique of PCR
 Application of PCR.

Technique of PCR
 Denaturation of DNA.
 Annealing with 2 primers.
 DNA amplification by
synthesis of new DNA
strand in the presence of
enzyme DNA polymerase
& substrate
deoxyribonucleotide
triphosphate.

Application of PCR.
 Rapid diagnosis of AIDS.
 DNA fingerprinting in kinship analysis &
identification of crime suspect.
 Prenatal diagnosis of genetic disease.
 Study of evolution from DNA of archeological
samples
 Sex identification.

BLOTTING TECHNIQUES.
 Analytical techniques
used for identification
of special DNA, RNA or a
protein.
 Southern blotting
 Northen blotting
 Western blotting.

SOUTHERN BLOTTING
 It is the name after the
scientist who identified
it.
 Applications
 DNA finger printing.
 Detection of mutant gene
causing cystic fibrosis.

NORTHEN BLOTTING
 Similar to southern
blotting except that it is
for RNA instead of DNA.
 Application
 Analysis of expression of
gene in a particular
tissue.

WESTERN BLOTTING.
 Technique for
identification of specific
protein.
 Application
 Confirmatory test for
HIV.
 With positive ELISA test
positive western blot is
99.9% accurate in
detecting HIV infection.

CLONING.
 Making of many
identical copies of a
molecule.

Some important terms
 Transformation – introduction of any DNA molecule into any
living cell.
 Transfection – introduction of purified DNA molecule into
cultured cell.
 Transduction – transfer of genetic material or DNA from one
cell to other with the help of virus.

Some important terms
 Microinjection – introducing new DNA into cell by injecting it
directly into nucleus.
 Biolistics – introducing DNA into cell that involves
bombardments with high velocity microprojectiles located
with DNA.

CLONING.
 Types
 Gene cloning
 Reproductive cloning.
 Embryo cloning.(therapeutic cloning)
 Tissue culture.

Gene cloning
 Refers to the process of transfer
of a DNA fragment of interest
from one organism to a self-
replicating genetic element
(cloning vector) such as
bacterial plasmid
 and subsequent propagation of
the recombinant DNA molecule
in the host organism

Applications of gene cloning, i.e.
recombinant DNA technology
 It include:
 Gene therapy,
 Gene engineering of organisms and
 Sequencing genomes.

Reproductive Cloning.
 Technology used to
generate an animal
that has same nuclear
DNA as another
currently or
previously existing
animal.

 In this technique genetic material is transferred
from the nucleus of a donor adult cell to an egg
whose nucleus, and thus its genetic material,has
been removed.

 A sheep (Dolly) was created
by somatic cell nuclear
transfer process by the
research team at the Roslin
Institute in Edinburgh
(Scotland) in Feb 1997.

Applications.
 In 2001, the first clone of an endangered animal, a
wild ox called a gaur was born. The young gaur
died from an infection about 48 h after its birth.
 In Italy (2001) a successful cloning of a healthy
baby mouflon, an endangered wild sheep was
reported.
 The cloned mouflon is living at a wild centre in
Sardinia.

Embryo cloning.(Therapeutic
cloning)
 Refers to the production
of human embryos for
use in research.
 Aimed at harvesting
stem cells that can be
used to study human
development and to treat
diseases rather than to
create a cloned human
being.

cloning)
 Stem cells are extracted from the egg at the
blastocysts stage of development and can be
used to generate virtually any type of
specialized cells in the human body.

cloning)
 First human embryo for the purpose of
therapeutic research was cloned in Nov,
2001 by the scientists from ‘Advanced Cell
Technologies (ACT), a biotech company in
Massachusetts.

Tissue culture.
 Certain cells when placed in a
suitable medium can be
cultured indefinitely.
 Allows a study of the action of
such chemicals as hormones,
drugs, antibiotics, cosmetics
and pharmaceutical products to
be made on cells.

APOPTOSIS.
 Programmed cell death,
occurs under genetic
control.
 Also called cell suicide.

Apoptosis for proper
development of tissue.
 In central nervous system -- The remodelling that
occurs during development and synapse formation.
 During formation of the fingers and toes of the
fetus, the apoptosis plays an important role of removing
the web tissue between the finger and toes.

Apoptosis for proper
development of tissue.
 During sexual development in fetal life the
apoptosis is responsible for regression of duct systems.

Apoptosis for normal
functioning of adult tissues
 Cyclic breakdown of endometrium that
leads to start of menstruation is caused by
apoptosis.
 Epithelial cells that lose their connection to
the basal lamina and surrounding cells
undergo apoptosis.
 Enterocytes sloughed off the tips of intestinal
villi undergo apoptosis.

Apoptosis to destroy the cells that
represent a threat to integrity of the
organism.
 Apoptosis of the cells infected with virus.
 Apoptosis of the cells of the immune system to
prevent them from attacking body
constituents.
 Apoptosis of the cells with DNA damage.
 Apoptosis of the cancer cells.

MECHNISM OF APOPTOSIS.
 The final common pathway. - activation of
a group of cysteine proteases called caspases
 which exist in cells as inactive proenzymes
 Triggering stimuli
 Internal stimuli
 Apoptosis’s inducing factor
 External stimuli.

MOLECULAR GENETICS &
MEDICINE.
 Mutations & genetic human disease
 Detecting human genetic variations
(genetic screening)
 Genetics & cancer
 Gene therapy.

MUTATIONS & GENETIC
HUMAN DISEASE
 Mutations
 Point mutations
 Frame shift mutations.
 Genetic human
diseases.

DETECTING HUMAN GENETIC
VARIATIONS (GENETIC SCREENING)
 Field of genetic screening.
 Prenatal diagnosis-
 Chorionic villus sampling,
 Amniocentesis and
 Pre-implantation diagnosis.

DETECTING HUMAN GENETIC
VARIATIONS (GENETIC SCREENING)
 Carrier diagnosis –
 Identification of people who carry a particular genetic
disease, usually with no visible symptom or harm to
themselves.
 Examples -- Sickle cell anaemia, cystic fibrosis and
phenylketonuria.
 Predictive diagnosis.
 Prediction of a future disease from which one is likely to suffer.
 Example -- 'genetic time bomb' is Huntington's chorea where the
onset of disease occurs in middle age.

GENETICS & CANCER
 Points favoring genetic basis for cancer –
 Hereditary predisposition is noted in some cancers
like colon cancer and retinoblastoma.
 Chromosomal abnormalities are noted in many
forms of cancers like Burkitt's lymphoma and acute
myeloid leukaemia.

GENETICS & CANCER
 Defective DNA repair mechanisms have been
associated with occurrence of cancers.
 Genetic damages (mutagenesis) by the action of
various agents like ionizing radiations, UV rays
are associated with occurrence of cancer.

GENES & MOLECULAR FACTORS INVOLVED
IN PATHOGENESIS OF CANCER.
 Oncogenes - genes are derived by somatic
mutations from the closely related proto-
oncogenes
 Tumor suppressor genes
 Mutator genes
 Telomeres in cancer

Genetic changes converting proto-
oncogenes into oncogenes
 Missense mutation, i.e. change in the amino acid
sequence of proto-oncogene protein converts it into
oncogene.
 Gene amplification, e.g. Myc genes have been amplified
in human leukaemia, breast, stomach, lung and colon cancer.

Genetic changes converting proto-
oncogenes into oncogenes
 Chromosomal translocations, e.g. in Burkitt’s
lymphoma a region of chromosome 8 is translocated to
either chromosome 2, 14 or 22. The breakpoint in
chromosome 8 causes the overexpression of c-myc gene.
 Retroviral integration, e.g. in avian lymphomas the
integration of the avian leucosis virus can enhance the
transcription of the c-myc gene.

GENE THERAPY.
 Methods that aim to
cure an inherited
disease by providing the
patient with a correct
copy of the defective
gene

GENE THERAPY.
 Basic principles.
 Gene replacement
 Gene correction.
 Gene Augmentation.

Basic Approaches
 Germ line therapy
 Somatic cell therapy
 Steps - involved in this therapy are:
 Isolation of the cells with the gene defect from a patient,
 Growing the isolated cells in culture,
 Transfecting the isolated cell with a remedial gene
construct,
 Selecting, growing and testing the transfecting cells and
 Either transplanting or transfusing the transfecting cells
back into the patients.

GENETICS

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