Genetic of Bacteria, Transformation, conjugation, Transduction

1. Genetics of Bacteria

2. Genetics of Bacteria
■ The genetic information of main bacteria is stored in a single main chromosome,
mini-chromosomes plasmids and episomes.
■ Main Chromosome: The chromosome carries a few thousand genes.
■ Plasmids: Plasmids are variable in number. Plasmids are autonomously
replicating, circular DNA that carry from three genes to several hundred genes.
Some bacteria contain as many as 11 different plasmids in addition to main
chromosome.
■ Episomes: Episomes are similar to plasmids. But Episomes can replicate
autonomously Or as part of the main chromosome in an integrated state.
Reproduction in bacteria
■ Bacteria produce asexually by simple fission. Each daughter cell receives one copy
of chromosome. The chromosome of bacteria do not go through mitotic and
meiotic condensation cycles. Therefore recombination like independent
assortment and meiotic crossing over do not occur in bacteria. Parasexual process
occur in bacteria.

3. Mutant Genes in bacteria
■ Bacteria will grow in liquid medium or on the surface of semisolid medium
containing agar. If grown on semisolid medium, each bacterium will divide and
grow exponentially. They produce a visible colony on the surface of the medium.
The number of colonies that appear on a culture plate can be used to estimate
number of bacteria. Each bacterial species produces colonies with a specific color
and morphology. Mutations in bacterial genes can change both colony color and
morphology.
■ Mutants blocked their ability to utilize specific energy sources. Wild-type E.coli can
use almost any sugar as an energy source. However, some mutants are unable to
grow on the milk sugar lactose. They grow well on other sugars but cannot grow on
medium containing lactose as the sole energy source. Other mutants are unable to
grow on galactose. Therefore, wild-type E.coli is lac+ (able to use lactose as).
■ Mutants that are unable to use lactose are lac-.

4. Unidirectional Gene Transfer in
Bacteria
■ Gene transfer is unidirectional rather than bidirectional.
Recombination events in bacteria occur between a fragment of one
chromosome (from a donor cell) and a complete chromosome (in a
recipient cell).
■ Crossovers must occur in pairs and must insert a segment of the
donor chromosome into the recipient chromosome. If a single
crossover occurs, it will destroy integrity of the recipient
chromosome, producing a nonviable Linear DNA molecule.

5. Recombination in bacteria

6. Mechanism of Genetic Exchange in
Bacteria
■ Bacteria transfer or receive genetic material in 3 different ways:
1. Transformation
2. Conjugation
3. Transduction
■ These three processes can be distinguished by two simple criteria
■ Does the process require cell contact?
■ Is the process sensitive to deoxyribonuclease (Dnase)? Dnase is an enzyme that
degrades DNA.

7. Mechanism of Genetic Exchange in
Bacteria

8. Transformation

9. Transformation
■ Transformation is the process by which a bacterium will take up extracellular DNA
released by dead bacterium.
Discovery of transformation
■ It was discovered by Frederick Griffith in 1928 while working with the strains of
streptococcus pneumoniae. There are two forms of streptococcus pneumoniae
bacterium:
1. Normal form or S-form(Smooth): Normal pathogenic form of this bacterium forms
smooth colonies on a culture dish. So, it is called S-form. They are virulent.
2. Mutant form or R-form(Rough): The mutant form lack an enzyme which
manufactures the polysaccharide coat. They form rough colonies. So, they are
called R-form. They are non virulent.

10. Experiment of Frederick Griffith
■ Griffith performed following experiments:
1. He infected the mice with S strain of streptococcus pneumoniae bacteria.
The mice died of blood poisoning.
2. He infected the similar mice with R strain of streptococcus pneumoniae.
This bacteria lack polysaccharide coat. So they are non virulent. The mice
did not become ill. Thus, its outer coat is necessary for virulence.
3. Griffith wanted to know whether the polysaccharide coat itself had a toxic
effect. He injected dead S strain of bacteria into the mice. The mice
remained perfectly healthy. It means coat itself has no toxic effect.
4. Now, he injected mice with a mixture containing dead S-bacteria and live
coatless R-bacteria. Both of these strain are themselves harmless. But the
mice develops the disease symptoms and many of them died. The blood
of mice contained large number of virulent S-bacteria. These bacteria had
surface proteins of live (previously R) strain.

11. Continue:
■ Thus, the information of
polysaccharide coat has
passed from dead S-
bacteria to live R-
bacteria in the mixture.
It permanently
transforms the coatless
R-bacteria into the
virulent S variety. It is
called transformation
(transfer of genetic
material from one cell
to another and altering
the genetic makeup of
recipient cell).

12. Experiment of Avery, MacLeod and
McCarty
The agent responsible for transforming streptococcus remained undiscovered until
1944. Oswald Avery and his colleagues performed series of experiments and called it
Transforming principle.
1. They first prepared mixture of dead S- Streptococcus and live R- Streptococ. Then
they removed the protein from this mixture. They achieved 99.98% purity. But
transforming activity was not reduced.
2. The properties of transforming principle resembled DNA. The protein digesting
enzyme or RNA digesting enzyme did not affect the transforming activity. But the
DNA digesting enzyme DNAase destroyed all transforming activity. So, they proved
that DNA is responsible for transforming principle.
3. The ability of bacteria to take up foreign DNA is called Competence.

13. Mechanism of transformation
DNA transfer in Bacteria by transformation involves following
steps:
1. dsDNA attaches to membrane bound dsDNA binding
protein
2. One of two strands of transforming DNA passes into the
cell while the other strand degraded by a nuclease
(deoxyribonuclease)
3. Single strand exogenotes are unstable and will usually be
degraded unless they are integrated into the endogenote.
By the process of homologous recombination the
transforming DNA integrates the bacterial chromosome.

15. Conjugation

16. Conjugation
Transfer of genetic material between bacteria through cell-to-cell contact is called
bacterial conjugation
Horizontal genetic transfer
Genetic material transferred by Sex F Factor
It doesn’t involve fusion of gametes and formation of zygote. Therefore, it is not
actually sexual mating. It is only transfer of genetic material from donor cell to
recipient cell. Mating types in bacteria are:
1. Donor cell contained F factor (Fertility Factor or Sex pillus) donates genetic
material.
2. Recipient cell doesn’t contain F Factor receives genetic material.
3. The Conjugate – HFR (High Frequency Recombinant), Resulting genetic
recombinant, transfer entire genomic DNA when mixed with recipient cell.

17. Culture of bacteria For conjugation
■ There are two culture of bacteria:
1. Some culture of cells contain non-integrated F-Plasmid.
These are responsible for low frequency of chromosomal
gene transfer.
2. Some strains of bacteria have Integrated F-Plasmid. Such
strains transfer chromosomal genes very efficiently. So they
are called HFR (High Frequency Recombination).
A HFR cell is a bacterium with a conjugative plasmid
(often F) integrated into its genomic DNA.

18. Mechanism of Conjugation
1. The prototype for conjugative plasmid is F- plasmid also called F-Factor. The F-
plasmid is an Episome. Episome is a plasmid if about 100kb length that can
integrate itself into the bacterial chromosome by genetic recombination. (1kb=1000
base pairs).
2. There can only be one copy of F-Plasmid in a bacterium. Such bacterium is called F-
positive or male bacterium. The bacteria lacking F-Plasmid is called F-negative or
female bacterium.
3. The F-Plasmid carries pilli gene and regulatory genes. These genes form sex pilli on
the cell surface. They also form polymeric proteins. These proteins can attach
themselves to surface of F-negative bacteria and initiates conjugation. F-Plasmid
also form some proteins which causes the opening of channel between bacteria.
4. Conjugation initiated by mating signal. A complex of protein Relaxosomes contain a
nick in one plasmid DNA strand. Relaxosomes contain many proteins and
integrating host factor (IHF).

19. Continue:
■ Mating always occur between
F+ and F-. F-Plasmid is
integrated into host genome.
Donor chormosomal DNA may
be transferred along with the
Plasmid DNA. The
chromosome of donor bacteria
replicated before this transfer.
Therefore its gene are not lost.
■ The amount of chromosomal
DNA transferred depends on
the timing of contact. For
common laboratory strains of
E.coli the transfer of entire
bacterial chromosome takes
about 100minutes. The
transferred DNA can be
integrated into the recipient
genome via recombination.

20. Transduction

21. Transduction
■ Transduction is the process by which bacterial DNA is moved from one
bacterium to another by a virus.
Discovery
■ It was first discovered by Joshua Lederberg in 1952. The most striking
feature is the transfer of genetic material from cell to cell by viruse.
The second feature is the fact that only a small part of the total
genetic material of any one bacterial cell is carried by any particular
transducing particle.

22. Mechanism of Transduction
1. Bacteriophage infect a bacterial cell. Their normal mode of reproduction is to use the
DNA replication machinery of the host bacterial cell. Thus this bacteriophage makes
numerous copies of their own DNA or RNA. These copies of bacteriophage DNA or RNA
are then Packaged into newly synthesized copies of bacteriophage virions.
2. However, packaging of Bacteriophage DNA is not fool-proof. Small pieces of bacterial
DNA also pack into a bacteriophage virion along with bacteriophage genome. Viruses
with RNA genome are not able to package DNA. Thus they don’t usually make this
mistake.
3. Theses virions come out on lysis of host cell. The mispackaged virions containing
bacterial DNA. It can attach to other bacterial cells. They inject the DNA they have
packaged. Thus they transfer bacterial DNA from one cell to another. This DNA can
become part of the new bacterium’s chromosome.

23. Types of Transduction
There are three types:
1. Generalized Transduction: It occurs in lytic cycle of phage virus. DNA of phage
virus enter into E.coli bacteria. This DNA replicates and develop many new DNA
and capsids. The DNA of bacteria is broken. Some pieces of DNA also enter into
capsid of viruse. Bacteria burst and release new phage viruses. Now this phage
enters into recipient bacteria and transfer DNA of donor bacteria into the DNA of
recipient bacteria. Bacterial endonuclease enzymes destroy the phage virus. Now
these bacteria incorporate genes of donor bacteria and replicates.
2. Specialized Transduction: It occurs in lysogenic cycle of phage viruse. In this cycle,
viral DNA incorporated into bacterial DNA as prophage. It remains peacefully
there. But sometimes, it becomes lytic. It comes out of bacterial DNA. Some part
of bacterial DNA remains attach with it. Viral DNA with a piece of bacterial DNA
replicates and develop new capsids. Bacteria burst. Virus infects other bacteria
and transfer genes of donor Bacteria to recipient bacteria.

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3. Restricted Transduction:
Certain phages carry out a more restricted kind of Transduction. They carry out only a
specific section of bacterial genetic material. They transduce only a few genes.
Retroviruses carry out specific or restricted Transduction. These viruses can cause the
formation of tumors (oncogenesis) in animals. It is now known that these viruses
exchange a small portion of their genome for a mutant cellular gene. It has a role in
gene regulation or replication. These viruses carrying mutant genes to infect cells.
They transform these cells into tumor cells.

26. Thank you😊