DNA molecule is composed of 2 strands of complementary nucleotides bound together by a double Helix. Bacterial nucleus contains circular chromosome of a double strand DNA molecule of 1000um (1mm) long when straightened. Each strand have a backbone of deoxyribose sugar and phosphate groups There are 4 nitrogenous bases Two purines- adenine(A) and guanine(G) Two Pyrimidines- thymine(T) and cytosine(C) One of these four nitrogenous bases is attached to each deoxyribose (sugar) The two stands are held together by hydrogen bonds between the nitrogenous bases on the opposite strands

1. BACTERIAL GENETICS
Dr Roma Goyal
Assistant Professor
Department of Microbiology

2. Understanding Genetics
We resemble and differ because of Genetic
configuration.
Parents- Son- Daughter
HOW THEY RESEMBLE EACH OTHER?
They breed true from Generation to Generation..
But vary in small proportion in progeny.
Bacteria too obeys the law of Genetics.

3. Watson and Crick
Discovery of DNA

4. 4

5. Basic Principles
• DNA molecule is composed of 2
strands of complementary
nucleotides bound together by a
double Helix.
• Bacterial nucleus contains circular
chromosome of a double strand DNA
molecule of 1000um (1mm) long
when straightened

6. • Each strand have a backbone of deoxyribose sugar
and phosphate groups
• There are 4 nitrogenous bases
• Two purines- adenine(A) and guanine(G)
• Two Pyrimidines- thymine(T) and cytosine(C)
• One of these four nitrogenous bases is attached to
each deoxyribose (sugar)
• The two stands are held together by hydrogen bonds
between the nitrogenous bases on the opposite
strands

7. Basic Principles contd…
• This bonding is in such a specific manner that
hydrogen bonds can only be formed between
adenine and Thymine (A-T) and between
guanine and cytosine (G-C)
• Adenine and thymine (A-T) one
complementary base pair
• Guanine and cytosine (G-C) form one
complementary base pair

8. Basic Principles contd…
• RNA is structurally similar to DNA
• Except it has sugar base ribose is present
instead of deoxyribose
• And the nitrogenous base uracil instead of
thymine
• There are three different types of RNA in a cell
• Messenger RNA (mRNA), ribosomal RNA
(rRNA) and transfer RNA (tRNA)

9. Important definitions
• Gene- A segment of DNA that specifies for a
particular polypeptide is called a gene.
• DNA contains many Genes( A combination of
hundreds and thousands of Nucleotides )

10. Codon
• Codon –Genetic information is stored in the DNA as a
code. Codon consists of three nucleotide bases i.e.
codon is triplet
• Each codon specifies production of a single amino
acid but more than one codon exist for the same
amino acid e.g. AGA codes for arginine but CGU,
CGC,CCG and CGA and AGG also codes for same
amino acid
• This is also applicable to other amino acids

11. Non-sense codon
• Three codons (UAA,UGA and UAG) do not
code for any amino acid and act as stop
codons for terminating the message for the
synthesis of a polypeptide
• These are called non-sense codons

12. Extra chromosomal genetic elements
• Some bacteria posses extra
chromosomal DNA this is
called as plasmids (When
situated in cytoplasm).
• Episomes When extra
chromosomal DNA is
integrated with
Chromosome of the
bacteria

13. Plasmids
• Plasmids are circular DNA molecule and can
replicate autonomously
• It is often not possible to differentiate
between plasmids and episomes
• Plamids and episome are not essential for
bacteria
• Plasmids carry properties of drug resistance ,
toxigenicity, conjugation and others

15. Plasmids contd…
• Some plasmids have the ability to transfer
themselves to other bacteria of the same or other
species. These are called self transmissible plasmids
• Transfer of plasmids occur by conjugation
• There are other plasmids which cannot be
transferred themselves (non transmissible) but can
be transduced
Ability of plasmids to transfer DNA from one cell to
another
• Plasmids have become important vectors in
recombinant DNA technology or genetic
engineering

16. Genetic variation
• Genotypic variation occur in the genetic
material.
• They are stable, heritable and not influenced
by environment
Genotypic variation occur by
• Mutation
• Gene transfer

17. Mutation
• Mutation is an inherited change in the nucleotide sequence
of the nucleic acid comprising the genome of an organism .
• A strain carrying such changes is called as mutant.
• A mutant may differ from its parent strain in genotype
(sequence of nucleotides in the DNA of the genome) and
sometimes in phenotype (observable properties from its
parent) also.

18. Mutation
• Mutation can be either spontaneous or induced.
• Spontaneous mutation occurs naturally (natural
radiation or due to error in pairing of bases during
replication).
• Mutation involving one or a very few base pairs are
referred to as point mutations.
• Mutation involving change in base pairs without
causing change in the amino acid that code for is
called silent mutation.

19. Mutation
• Mutation involving change in base pair which codes for a
different amino acid is called missense mutation. Eg. (UAC -
Tyrosin; AAC– asparagine).
• Some times a mutation may result in premature termination
of translation (as the base pair alteration contribute to stop
codon TAG - UAG (stop codon) resulting in incomplete protein
– such is called non-sense mutation.

20. 20
Mutation
Mutations can arise spontaneously in bacteria
-Also caused by radiation and chemicals
Mutations (and plasmids) can spread rapidly in a
population
-Negative consequences for humans
-For example:
-Methicillin-resistance Staphylococcus aureus (MRSA)
-Vancomycin-resistant Staphylococcus aureus (VRSA)

21. Transmission of Genetic material
(Gene transfer)
Prokaryotes do not reproduce sexually
However, they undergo horizontal gene transfer.
Various methods are
• Transformation (uptake of naked DNA)
• Transduction (Through bacteriophage)
Lysogenic conversion
• Conjugation (Plasmid mediated)

22. 22
TRANSFORMATION
Transformation
-It is the transfer of genetic information through
free or naked DNA
-DNA that is released from a dead cell is picked
up by another live cell
Occurs in many bacterial species, including
Streptococcus which was studied by Griffith

23. The Griffith’s Experiment
• In 1928, Frederick Griffith discovered
Tranformation while working on Streptococcus
pneumoniae.
• The bacterium exists in two strains
– S
• Forms smooth colonies in culture plate
• Cells produce a polysaccharide capsule and cause disease
– R
• forms rough colonies in culture plate
• Cells do not produce a polysaccharide capsule and are
harmless.

24. Transformation
• Genes are transferred from one bacterium to
another as “naked” DNA
• Frederick Griffith (1928)

25. 25
TRANSDUCTION
- Transmission of a portion of DNA from one
bacterium to another through the BACTERIOPHAGE
• In this bacteriophage nucleic acid and a portion of
the host DNA may be accidentally incorporated into
the bacteriophage. This is known as packaging error
• This bacteriophage when infects another bacterium
the host DNA transfers and the recipient cell
acquires new characters coded by donor DNA

26. Viruses infecting a bacterium

27. Bacteriophage have two types of
life cycle inside the host
bacterium
• Virulent or lytic cycle
• Temperate or non lytic cycle

28. Virulent or Lytic cycle
• In this large number of progeny is formed and
subsequently these progeny phages are
released causing death and lysis of the host
cell

29. Temperate or nonlytic cycle
• In this the host bacterium remains unharmed The
phage DNA (new genetic element) remains
integrated with the bacterial chromosome as
prophage which multiplies synchronously with
bacterial DNA.
• The prophage act as additional chromosomal
element which encodes for new characters and
transferred to daughter cells
• This process is known as lysogenic conversion and
bacteria harboring prophage are known as lysogenic
bacteria

31. 31
CONJUGATION
• The transfer of genetic material from one bacterium
(donor male) to another bacterium (recipient female)
by contact or mating is called conjugation
• Donors bacteria are those bacteria that contain F
plasmid ( F+ male cells) while cell lacking F plasmid is
called (F- or female cells )act as recipients
• F plasmid is conjugative plasmid which encodes for sex
pilus (F+ cells) which is necessary for conjugation . This
plasmid is known as sex factor or fertility factor

32. 32
CONJUGATION
• During conjugation the plasmid DNA replicates and
copy of it passes from donor to the recipient cell
probably along the sex pilus (conjugation tube) as a
result recipient (F-) becomes F+ donor (F+) and can
in turn conjugate with other female cells (F-)
• This character of maleness (F+) in bacteria is
transmissible or infectious

33. Sexduction
• F plasmid has the ability to
integrate into its own host
chromosomes.
• These cells transfer chromosomal
DNA to recipient cells with high
frequency (Hfr cells)
• Conjugation with Hfr cell and f-
rarely becomes F+ cell but it
receives chromosomal DNA from
donor

34. Transposable Genetic elements
• Transposable Genetic elements are specific
sequences of DNA segments that have the ability to
move from one plasmid to another plasmid or from
plasmid to chromosome and visa versa and also with
in the chromosome
• Because of their ability to insert into many sites both
on plasmid and chromosome they are called jumping
genes
• The transfer of genetic material from one DNA
molecule to another is called transposition

35. Detection methods of Genetic material
(DNA/RNA )
• Polymerase chain reaction(PCR)
• DNA probes
• Blotting techniques.

36. POLYMERASE CHAIN REACTION (PCR)
• Kary Mullis (1993) awarded Noble prize for his
discovery.
• Nucleic acid amplification system
• Produces large amount of DNA.
• Amplifies specific DNA sequence. (gene)

37. • PCR involves four main stages.
–Denaturation
–Primer annealing
–DNA synthesis
–Detection of amplified
product.

38. POLYMERASE CHAIN REACTION (PCR)
STEP-1
• Denaturation
 The double stranded DNA is dislocated to
single stranded DNA.
 Denaturating temperature (94ᵒC)

39. POLYMERASE CHAIN REACTION (PCR)
STEP-2
• Annealing of Primers
 Oligonucloetide primer attaches to target
DNA.
 Temperature is reduced to 50-60 ᵒC
(Annealing temperature).

40. POLYMERASE CHAIN REACTION (PCR)
STEP-3
• DNA synthesis
 Polymerase enzyme derived from Thermus
aquatics(Taq) triggers the formation of new
DNA strand.
 Repeat these three steps till 20-30 cycles.
 This is automated process done in
thermocycler/PCR machine.
 Exponational increase in the amount of DNA
occurs.

41. STEP-4
• Detection of amplified product
 Amplified DNA can be detected by Gel
electrophoresis.

43. POLYMERASE CHAIN REACTION (PCR)
Types of PCR:
–Reverse transcriptase PCR (RT-PCR)
–Nested PCR
–Multiplex PCR
–Real Time PCR

44. Application of PCR
• PCR provides a rapid tool for diagnosis of
various diseases be their infectious,
Neoplastic, or Genetic or in Forensic
investigations
• In this a specific DNA sequence of a particular
infectious agent is amplified with specific
primers

45. POLYMERASE CHAIN REACTION (PCR)
PCR helps in diagnosis of infectious disease:
• Bacterial infections: M. tuberculosis, Legionella pneumophilia, H.
pylori, C Trachomatis, Mycoplasma pneumoniae
• Viral infections: CMV, Herpes simplex, Hepatitis B, C, Coxsackie
virus, measles virus, HIV I & 2,HPV,Rotavirus,Rubella virus,
Adenovirus, parvovirus
• Fungal infections: C. Albicans, Cryptococcus, Pneumocystis jiroveci,
• Parasitic infections: T. Gondii, Plasmodium, Trypnosoma

46. DNA Probes
• DNA probe A single-strand DNA fragment used to
detect the complementary fragment.
• DNA probes are used widely in
bacteriology/Molecular diagnosis.
• Recombinant DNA techniques are used to isolate,
reproduce, and label a portion of the genetic
material, DNA, from the nucleus of a microorganism
that is specific for it.
• This fragment can be added to a specimen containing
the organisms.

47. DNA Probes contd…
• The specimen and known DNA are treated so that
the DNA strands from the organisms in the specimen
are separated into single strands.
• The DNA from the specimen rejoins (is annealed to)
the known labeled DNA and is thereby labeled.
• This permits the identification of a single pathogenic
organism in a specimen that contains many different
microorganisms.

48. DNA Probe

49. Application of DNA probes
• In clinical microbiology
Direct detection of microorganisms
to detect microbes difficult to culture
• Identification of culture isolates
• Strain identification
• To identify toxins, virulence factors
• Identification of resistant markers

50. Blotting Techniques
blotting method A technique for analysing a
tiny portion of the primary structure of
genomic material (DNA or RNA).
• Southern blotting method
• Northern blotting method
• Western blotting method

51. Southern blotting method
• Southern blotting method A technique used in
molecular genetics to analyze a small portion of
DNA.
• First by purifying it, then by controlled
fragmentation, electrophoretic separation, and fixing
the fragment identity using specific DNA probes.

52. Northern blotting method
• Northern blotting method A blot analysis
technique for analyzing a small portion of
RNA.
• Operationally, this test is identical to
Southern blotting except for the target (RNA)
and the specific reagents used.

53. Western blotting method
• Western blotting method A technique for
analyzing protein antigens.
• HIV proteins

54. Genetically modified organisms
• The process of artificially introducing foreign DNA
into organisms is called Transfection
• The recombinant organism produced in this way is
called transgenic or genetically modified organisms
• Transgenic organisms are available for a variety of
biotechnological applications

55. Gene Therapy
• It is a therapy by which faulty gene is replaced
by the normal gene in persons suffering with
fatal or debilitating disease
• Main benefit is defective gene is replaced with
normal gene
Types
• Ex Vivo therapy
• In Vivo therapy

56. Gene Therapy contd…
Ex Vivo Therapy In this normal gene is cloned
with vectors like adenoviruses which are
infectious but harmless .
• Tissues are removed from patient and
incubated with these genetically modified
viruses to transfect them with normal gene
• The transfected cells are reintroduced in the
patient by Transfusion

57. Gene Therapy contd…
In Vivo Gene Therapy normal gene is cloned
with vector as adenovirus and it is then
directly introduced into the patient tissues

58. Applications of Gene Therapy
• A person born with defective genes can have
is defective gene replaced by this method
• A cancer patient who have developed
defective gene which have caused the cancer
can have his defective gene replaced and can
lead a normal life

59. Thank You !!!