Bacterial genetics

Concepts of
Bacterial
Genetics
DR. Mishuk Shaha
DVM (CVASU), MS (Animal Breeding
and Genetics)

*
*Genetic material inside a bacterium is often called:
Nucleoid/Nuclear body/bacterial nucleus
*Chemical composition of nucleoid of bacteria (E. coli
and others)
*About 80% DNA
*10% RNA (mostly nascent)
*10% protein (mostly RNA polymerase)
DR. Mishuk Shaha

*DNA molecule is supercoiled, and this accounts for its
efficient packing inside the cell.
*In order for the double stranded helical DNA molecule to
be supercoiled, one strand must first be broken (nicked)
so that the helical molecule can be twisted upon itself
(supercoiled).
*Studies suggest that there are 18 to 20 loops of DNA in
the E. coli chromosome and that each loop is
supercoilded, so that the entire molecule is reasonably
compact. (Chromosome’s length of a bacterium may be
1000 times longer than the entire cell length)
DR. Mishuk Shaha

Genetic Elements of bacteria:
* Genome = Complete set of genetic elements occurring
within a cell.
In case of bacteria, Genome = genetic information carried
by the Chromosome and the Extra chromosomal plasmids
* A plasmid is an extra-chromosomal DNA molecule separate
from the chromosomal DNA which is capable of replicating
independently of the chromosomal DNA. In many cases, it is
circular and double-stranded. Plasmids usually occur
naturally in bacteria, but are sometimes found in eukaryotic
organisms (e.g., the 2-micrometre-ring in Saccharomyces
cerevisiae).
* The term plasmid was first introduced by the American
molecular biologist Joshua Lederberg in 1952.
DR. Mishuk Shaha

* Plasmid size varies from 1 to over 1000 kilobase
pairs(kbp).
* The number of identical plasmids within a single cell
can range anywhere from one to even thousands
under some circumstances.
* Plasmids can be considered to be part of the
mobilome, since they are often associated with
conjugation, a mechanism of horizontal gene transfer.
* Similar to viruses, plasmids are not considered a form
of "life" as it is currently defined.
DR. Mishuk Shaha

*Most nongrowing eukaryotic cells are diploid, that is, they
contain two copies of each chromosome per cell (pairs of
matching genes).
*Eukaryotic chromosomes are structurally complex, and cells
have a number of different types of chromosomes.
*In contrast with eucaryotes, bacteria have simple genetic
systems.
*Bacterial chromosomes are single DNA molecules.
*Non-growing cells are haploid; that is, they contain only one
chromosome(DNA molecule) per cell (i.e., one set of genes).
*During growth, however, the bacterial cells contain at least
one partial copy of its chromosome at any one time, because
DNA synthesis must provide two complete copies just prior to
cell division.
DR. Mishuk Shaha

*
Genetic processes require at least three types of polymers:
*Deoxyribonucleic acid (DNA)
*Ribonucleic acid (RNA)
*Proteins
*During cell growth, all three types of macromolecules are made.
* DNA and RNA are often called informational macromolecules to
distinguish them from others.
*The cell’s structural genes determine the order of placement for
each amino acid during the synthesis of a protein molecule.
DR. Mishuk Shaha

*Gene: A gene is a molecular unit of heredity of a
living organism.
*Genes composed of nucleic acids
*Information stored in nucleotide sequence (A,T,G,C)
*Information used to assemble amino acids into proteins
*Proteins perform variety of essential cellular functions
*One gene consists of a sequence of triplet purine and
pyrimidine bases of the DNA molecule that codes for a
sequence of amino acids as the protein molecule is being
made.
*The base sequence in each triplet is critical, because a
change in even one base may cause an inactive protein to be
formed or actually stop the synthesis of that protein.
DR. Mishuk Shaha

*DNA does not function directly in protein synthesis, however,
but through an intermediate molecule called mRNA.
*During mRNA synthesis, one strand of the DNA serves as a
template, so that the purine and pyrimidine bases on the
mRNA are assembled in the proper order during synthesis. The
triplet bases on the mRNA are called codons.
*The process of transferring information from DNA to an mRNA
molecule during synthesis is called transcription.
*Once mRNA synthesis is complete then the new molecule
binds to a ribosome and it serves as a template for protein
synthesis.
DR. Mishuk Shaha

*Each tRNA has two critical sites:
- One site will bind with one type of amino acid, and
* - the other site (on opposite side of the molecule) consists
of three purine and pyrimidine bases that are complementary
to only one codon on the mRNA
*The triplets of bases in the tRNA complementary to those on
mRNA are called anticodons.
*Each tRNA molecule binds to an amino acid and also binds to
the mRNA, so that the amino acid will be inserted in the
proper order during protein biosynthesis.
*This process of transferring the codes found in the mRNA into
the sequence of amino acids in a protein is called
translation.
DR. Mishuk Shaha

DNA Structure and Synthesis
*A DNA molecule consists of two strands, each of which
contains alternating units of phosphate (HPo4
2-
) and a
sugar called deoxyribose.
*Each phosphate attaches to both the 3′ and 5′ position of
adjacent deoxyribose molecules (by ester linkage),
forming what is called’suger-phosphate backbone’.
*At one end of each strand, the sugar phosphate has a
free 3′ hydroxyl, and it has a free 5′ hydroxyl at the other
end. One of four purine and pyrimidine basess is attached
to each deoxyribose on each strand.
*Replication always begins at the 5′ end and progresses
toward the 3′ end of each DNA strand.
DR. Mishuk Shaha

*Two complementary strands of DNA are found in a
helical fashion (see the figure).
*With the ends the two strands covalently bonded
together to form a circular macromolecule.
DR. Mishuk Shaha

*How DNA replicates?
by unwinding the existing helix and then adding sugar-
phosphate-base units (called nucleotides). The point at
which the two strands unwind is called-the replication
fork or the growing point or the replicon
*It is believed that the bacterial chromosome is attached to
the cell’s plasma membrane at the DNA replication site.
*Each nucleotide unit is added at the 3′ hydroxyl end of the
new DNA strand. Therefore, replication of these two strands
runs in opposite directions.
DR. Mishuk Shaha

*When the double helix opens up to begin replication, an
enzyme called primase initiates the synthesis of a short DNA
(primer) strand.
*Once the first nucleotide is in place, then another enzyme
(one type of DNA polymerase) continues to covalently bond
each additional nucleotide to the 3′ hydroxyl end of the newly
developing strand. This new DNA forms in short chains (called
Okazaki fragments).
*The adjacent Okazaki fragments are covalently bonded
together by an enzyme called DNA ligase.
DR. Mishuk Shaha

*With the addition of each new nucleotide, the old DNA strand
acts as a template for the formation of the new strand, so
that complementary base pairing occurs during this synthesis.
*The end result of this synthesis is two double stranded DNA
molecules (two bacterial chromosomes).Each one contains
one strand from the old molecule and a new complementary
strand (that’s why called semiconservative).
*Once replicated, each chromosome is then supercoiled by the
enzyme called topoisomerases.
*The topoisomerase that promotes DNA super coiling (and thus
controls unwinding) at the replication fork is called DNA
gyrase.
DR. Mishuk Shaha

Speed of Replication
*Bacteria:
- The single molecule of DNA that is the E.
coli genome contains 4.7 x 106
nucleotide pairs.
- DNA replication begins at a single, fixed location
in this molecule, the replication origin, proceeds at
about 1000 nucleotides per second, and thus is done
in no more than 40 minutes.
- In bacteria, which have a single origin of
replication on their circular chromosome, this
process eventually creates a “ theta structure”
(resembling the Greek letter theta: ɵ)
DR. Mishuk Shaha

Enzymes involved in DNA replication of bacteria
*Primase-involves in the synthesis of primer
*DNA polymerase - regulates addition of nucleotide at3'end in
sequence to form Okazaki fragments
*DNA ligase - binds the adjacent Okazaki fragments together
*Topoisomerase - regulates supercoiling of DNA
*DNA gyrase - controls supercoiling and thus unwinding of DNA
at the site of replicon
DR. Mishuk Shaha

*
Three basic differences with DNA:
* -Possesion of ribose instead of the sugar deoxyribose
* -Uracil replaced the base thymine
* -Single stranded
DR. Mishuk Shaha

*Types of bacterial RNA
1. mRNA (messenger RNA)
2. t RNA (transfer RNA)
3. rRNA (ribosomal RNA
DR. Mishuk Shaha

* Messenger RNA (mRNA)
*Function of mRNA -The function of mRNA is to
copy (transcribe) the genetic code from the
gene (chromosomal DNA) and to move that
message to the site of protein synthesis (i.e.
the ribosome)
DR. Mishuk Shaha

mRNA synthesis
*mRNA synthesis begins from the Promoter region (the
site on the chromosome where mRNA synthesis begins).
Each gene or a set of genes has its own Promoter region.
*The enzyme RNA polymerase binds to the promoter
region. This binding initiates uncoiling of the two DNA
strands, so that the code may be read (transcribed)
from one DNA strand (called the sense strand).
DR. Mishuk Shaha

*
*The RNA polymerase then moves along the DNA sense
strand while it simultaneously bonds together the
ribonucleotide building blocks (ATP/CTP/GTP/UTP) to
form the new mRNA molecule.
*At the end of transcription the opened-up DNA helix
closes.
*Termination of mRNA synthesis also occurs at a specific
(termination) site on the DNA molecule.
DR. Mishuk Shaha

* t RNA (transfer RNA)
tRNA structure and function
* A tRNA is a single stranded molecule that contains
double stranded regions as a result of the molecular
folding back upon itself. tRNA looks like a clover leaf
with three distinct loops and the stem having two
open ends.
Functions:
* D-loop: selectively binds to an enzyme called an
aminoacyl-tRNA (AA-tRNA) synthetase. This enzyme
does two things :
(1) it specifically activates one type of amino acid; and
(2) it binds that activated amino acid to the acceptor
stem of the tRNA in the form of amino acid-AMP
complex.
DR. Mishuk Shaha

*The stem: As mentioned, binds with activated amino
acid.
*Anticodon loop: possesses three bases complementary
to a codon on mRNA to recognize the codon so that
specific amino acid can be inserted at the point in the
developing protein.
*TφC loop: appears to bind to the 50S ribosomal subunit
during the translation process to help the components
staying in the proper configuration while the activated
amino acids are being covalently bonded onto the newly
developing protein molecule.
DR. Mishuk Shaha

Structure of tRNA
DR. Mishuk Shaha

Function of tRNA
DR. Mishuk Shaha

*Ribosome structure and function
*Each ribosome has two subunits:
30S and 50S (S stands for Svedberg units, which is
based on sedimentation properties of the molecule in a
high speed centrifuge):
- Made up number of individual proteins and rRNA
- Each of the units exists separately
- When they come together in combination of a
molecule of mRNA they form 70S, which complex
helps in:
- translation of the genetic codes
- supporting the tRNA and many other proteins in
the translation process
*Distortion of ribosomal structure can prevent proper
functioning of the entire translational complex.
DR. Mishuk Shaha

*The Genetic Codes- are located in the DNA
*They are transcribed in the mRNA as codons; each codon
carries the information for the insertion of a single
amino acid during the process of phenotypic expression
of a gene.
*64 codons are possible for 20 amino acids (there are
only four bases in these combination and three bases
form a codon). Therefore, several codons are capable
for coding for insertion of the same amino acid.
(Example-UCU, UCC, UCG and UCA code for serine).
*A few codons, like UGA,UAG,UAA do not code for any
amino acid and they are called nonsense codons/Stop
codons. These codons serve as a signal to stop protein
synthesis when the protein molecule is fully formed.
DR. Mishuk Shaha

Mechanisms of protein synthesis in bacteria
Four steps involved:
*Initiation
*Elongation
*Termination
*Polypeptide folding
DR. Mishuk Shaha

1. Initiation
requires the formation of a complex containing mRNA
,ribosomal subunits, activated tRNA and initiation
factors (several proteins).
* In bacteria the first initiating codon is either AUG or
GUG. When used as an initiating codon, these triplets
code for an amino acid called N-formylmethionine, so
this is the first amino acid in the sequence for each
protein. This amino acid can later be enzymatically
removed, so not all bacterial proteins have
methionine at their amino terminal ends.
* (Initiation and amino acid activation require energy
which is supplied by ATP or GTP)
DR. Mishuk Shaha

2. Elongation: A repeated series of events:
(a) Recognition:A continual process where anticodons
matching with codons on the mRNA will be
recognized.
(b) Transferring of Amino acids: Once the two amino
acids are adjacent to one another, then the first
amino acid is transferred and covalently bonded to
the second amino acid.
(c) Translocation: The empty tRNA on the first codon is
released from the mRNA and the tRNA with its
attached polypeptide chain is moved along the
ribosome to the next position. This series of events is
repeated over and over again resulting in the
elongation of the polypeptide chain.
DR. Mishuk Shaha

3. Termination:
* When an mRNA codon is reached that does not code
for the attachment of any AA-tRNA, it indicates the
termination of the protein synthesis.
4. Polypeptide folding:
Once released from the ribosome-mRNA complex, the
polypeptide chain is free to fold into an active three-
dimensional protein structure that is held in this form
by weak disulfide bridges and hydrogen bonding.
DR. Mishuk Shaha

Mutation of bacteria
Sudden and inheritable change in the cell’s chromosome
is called mutation.
*Spontaneous mutation: is one that occurs without
known cause. Spontaneous mutations are probably
resulted most commonly from errors made while copying
the DNA during chromosome replication. For any given
gene spontaneous mutation may occur at the frequency
of only about one in every 105 to109 cells.
*Induced/Artificial mutation: though mutation occurs
spontaneously, its frequency can be increased markedly
(10-100 folds) by a number of agents collectively known
as mutagens.
DR. Mishuk Shaha

*Physical mutagens:X-rays, γ-rays, ultra violet light
*Chemical mutagens-Ethyl methane sulfonate, N-
methyl-N-nitro-N-nitrosoguanidine, nitrous acid,
acridine dyes etc.
*Molecular Mechanisms of mutagenesis
Mutations are resulted from alteration in the nucleotide
base sequence of the cell’s genes. There are two
mechanisms:
Point mutations
Deletion mutations
*Point mutations:
Resulted from the substitution of bases during DNA
replication-one deoxyribonucleotide is substituted for
another. Example: A deoxyadenosine may be inserted in
place of a deoxyguanosine.
DR. Mishuk Shaha

*Protein expression of point mutation depends on which
base has been substituted and where the substitution
has taken place.
*If there is no change takes place in the insertion of
amino acid (as there could be several triplets code for a
single amino acid), no change in the phenotypic
expression of this gene occur. This type of mutation is
called ‘Silent mutation’
DR. Mishuk Shaha

*Alternatively, the DNA -base substitution resulting from
point mutation can alter the triplet such that the mRNA
will code for the insertion of another amino acid during
protein synthesis.
*If this different amino acid is in a critical location, such
as part of an enzyme’s active site, then the function of
that protein could be altered or even destroyed,
resulting in a phenotypic change.
*If that protein were critical to the cell’s survival, this
point mutation could be lethal (Missense Mutation).
DR. Mishuk Shaha

*On the other hand, if the substitution altered the amino
acid sequence at a point on the protein not critical for
its catalytic activity, then this mutation would not be
phenotypically expressed.
*However, this mutation is not a silent one because the
amino acid sequence of the protein produced by that
gene is altered.
DR. Mishuk Shaha

Deletion mutation:
A portion of one strand of the DNA is removed (one to
several hundreds nucleotides may be deleted).
*Deletion of a single base in a structural gene will probably
result in a reading frame shift, and the next three bases may
be read as nonsense codon. This would cause the cessation
of transcription and hence a shorter mRNA formation.
*Deletion of a base at the terminal end of a gene, the
alteration of the mRNA and the resulting protein may not
drastically affect the protein’s catalytic activity. If, however,
the deletion occurs closer to the promoter end (the 5′end) of
the structural gene, then the resulting polypeptide will
probably be nonfunctional. Deletion of larger segments of
the DNA usually results in complete loss of the ability to
produce the protein.
DR. Mishuk Shaha

Reversion:
*Restoring the original characteristic that has been lost
due to mutation.
*Reversions = back mutations
*Reversion can be accomplished by using both physical
and chemical mutagens.
*(A good correlation exists between the mutagenic
capacity of a chemical and its carcinogenic ability.)
DR. Mishuk Shaha

Genetic recombination
Sudden and inheritable change caused by the introduction of new
genetic material from outside the cell.
*Ways/kinds of genetic recombination occur in bacteria
1) Transformation: when free chromosomal or plasmid DNA is
inserted directly into another recipient cell.
2) Transduction: Bacteriophage/phage (bacterial viruses)
mediated genetic recombination.
3) Conjugation: Bacterial mating that occurs after actual cell to cell
contact between a donor and a recipient cell.
*Transformation
To accomplish transformation two things are essential:
- an appropriate source of free DNA (from a donor
strain)
- competent recipient cells which are capable of:
- binding the foreign DNA molecules
- translocating the DNA across the cell
wall and plasma membrane
- inserting the DNA into its own chromosome
DR. Mishuk Shaha

Nature of Transformable DNA:
*The long continuously closed, supercoiled, helix of double
stranded DNA that serves as the chromosome within the
bacterial cell does not stay in that form when the cell is
lysed.
*Even with gentle lysis under laboratory conditions, the
chromosome will break into 100 or more pieces, each
piece containing about 50genes.
*A competent cell will usually incorporate only a few of
these DNA fragments, so that only a small portion of
genes from a donor cell can be transferred to another cell
by transformation.
Natural Competence:
*Unaltered cells that can take up free DNA fragments and
be genetically altered are said to be naturally competent.
DR. Mishuk Shaha

Artificial competence:
*It is possible to force some cells to become competent
by treating them with high concentrations of calcium
ions and subjecting them to temperature shocks to
increase the permeability of their cell wall and the
plasma membrane.
DR. Mishuk Shaha

*List of some bacteria with which natural transformation
has been described:
Gram- positive baceria
*Streptococcus pneumoniae
*Streptococcus sanguis
*Bacillus subtilis
*Bacillus cereus
*Bacillus stearothermophilus
Gram-negative bacteria-species of several genera:
*Neisseria
*Acinetobacter
*Moraxella
*Haemophilus
*Pseudomonas
DR. Mishuk Shaha

Steps involved in transformation:
1. Uptake of donor DNA by competent cells:
* The dsDNA first binds to specific proteins on the surface
of competent cells.
* In the case of the Gram-positive bacterium
Streptococcus pneumoniae,there are about 30 to 80
sites per cell, and only dsDNA will bind to these sites.
* Shortly after binding, one of the strands is degraded by
a cell-surface bound enzyme.
* Next, the resulting ssDNA is coated by a single, small
molecular weight polypeptide, and this complex enters
the cell by an unknown mechanism.
* In the case of Gram-negative Haemophilus and
Neisseria species studied, dsDNA is not degraded to
ssDNA before it enters the cell.
DR. Mishuk Shaha

2. Integration of Donor ssDNA into the chromosome
* Once inside the cell, the donor ssDNA from
Streptococcus pneumoniae tries to pair with a similar
region on the host cell’s chromosome.
* When a similar region is found:
a) one strand of the host’s dsDNA is opened up with an
enzyme called an endonuclease.
b) the dsDNA is unwound for a short distance.
c) the opposite end of the host DNA (corresponding to
the opposite end of the new donor DNA) is also cut
with an endonuclease.
d) the new donor strand is inserted and
e) enzymes called DNA ligases fuse both ends of the
donor ssDNA with the adjacent host chromosomal
DNA strand (Natural transformation is quite low as it
occurs from only 0.1to 1.0%).
DR. Mishuk Shaha

*In the case of gram-negative Haemophilus and Neisseria
species studied, the dsDNA that enters the cell is closely
associated with the host cell’s chromosome before one
strand is degraded.
*No free ssDNA intermediates seem to exist within the cell
prior to incorporation of the donor DNA into the host
chromosome.
*No single general mechanism appears to account for the
way in which the transformed DNA is incorporated into
the host chromosome in all bacteria.
DR. Mishuk Shaha

Artificial transformation can be achieved
by treating one of the following means:
*the calcium ions and cold-shock treatment
*Electroporation and
*sometimes, freez- thaw treatment and treatment of
protoplasts with polyethylene glycol
Escherichia coli are nor naturally competent cells but they
can be converted artificially in to recipient cells, and
there seems to be no trouble in getting the linear pieces
of ssDNA or dsDNA into the cell.
DR. Mishuk Shaha

*The problem is that, when the DNA molecules
are inside the cell they are quickly destroyed by
the host cell’s own nucleases before they can
be integrated into the chromosome.
*On the other hand, self-replicating forms of
covalently closed strands of DNA, like plasmids
and viral genomes are not attacked by the
recipient cell’s intracellular nucleases.
*These DNA can be inserted by force into the
normally incompetent cell. In this process, the
frequency of transformants in the survivors is
high (about 20%).
DR. Mishuk Shaha

*The process by which self-replicating forms of DNA are
artificially introduced into a cell that would normally not
be an appropriate host, and the expression of these new
genes in the recipient cell, are essential to the field now
called” genetic engineering”.
DR. Mishuk Shaha

Conjugation
*The conjugation is a process of bacterial mating where
DNA (mostly those of plasmids) transfer is mediated by
thin protein structure called sex pili (in case of gram-
negative bacteria) produced by the donor cells.
*Several sex pili are formed by the donor cell.
*It appears that the distal end of the sex pilus on the donor
makes contact with specific receptor sites of an
appropriate recipient cell.
*This process is followed by retraction which allows the
cells to come together until a conjugation bridge is
formed, through which the DNA is passed from the donor
to the recipient cell.
DR. Mishuk Shaha

*Mostly plasmid DNA is transferred through the process of
conjugation.
*Many plasmids contain transfer genes which lead to production
of sex pili.
*A single stranded copy of the plasmid DNA is transferred to a
recipient cell where a complementary strand is formed and
replication, as usual, occurs there after.
DR. Mishuk Shaha

There are three types of plasmids transferred in
the process of conjugation. These are:
*Fertility or ‘F’ plasmid
*Colicinogenic or ‘Col’ plasmid
*Drug resistance or ‘R’ plasmid
Fertility (F) plasmids:
*the first plasmid discovered in a strain of E coli.
*When F is in the cytoplasm of a cell it is called ‘F+’ or
male cell and a bacterium lacking the F plasmid is called
F- or female cell. F is readily transferred from the male
to the female by conjugation and the later becomes an F+
cell.
DR. Mishuk Shaha

Fig: The process of conjugation between an F+ and an F- bacteria
DR. Mishuk Shaha

*The F plasmid may become integrated in the bacterial
chromosome (a rare event) resulting in a ‘Hfr’ cell (High
Frequency Recombinations).
*Mating between Hfr cells and F- cells results in transfer
of part of the F genome plus some host cell genes from
the donor.
*Recipient F- cells usually remain F- following
conjugation because only part of the F plasmid from the
donor Hfr cell is transferred to the recipient cell during
the conjugative process.
*Therefore, these recipient cells will not possess the full
complement of genes that are subsequently necessary to
synthesize the sex pilus.
DR. Mishuk Shaha

*Recombination between the genetic material from the
donor cell and homologous regions in the F- recipient
enables the donor DNA to become expressed in the
recipient cell.
*The donor cell remains as Hfr because the host cell
(containing the integrated F plasmid) is replicated
during the transfer of the genomic single-stranded DNA
from the Hfr to the F- cell through the sex pilus.
DR. Mishuk Shaha

*Gram-positive bacteria are also able to exchange
genetic material via a conjugative process.
*but the transfer is not accomplished via a pilus, but
rather by a co-aggregation of the organisms in response
to production of pheromones by the donor bacterium.
*Under stimulation by these pheromones, potential
recipient bacteria synthesize a receptor molecule that
is specific for a conjugative adhesin present on the
donor cell.
*Aggregation results in the establishment of the cell-to-
cell connections that is necessary for mobilization of
the plasmid.
DR. Mishuk Shaha

Colicinogenic (Col) plasmids
*Strains of E.coli produce bacterial substances called
colicins which are active against other strains of E. coli
and closely related species.
*Similar substances are also produced by other bacteria
and are generally referred to as bacteriosins.
*The capacity for colicinogeny is transmitted by plasmids
during conjugation.
DR. Mishuk Shaha

Drug resistance transfer (R) plasmids
*Plasmids also carry genes that enable bacteria to grow
in the presence of one or more antimicrobial drugs.
*These plasmids are called ‘R’ plasmids or R factors,
which are transferred from cell to cell either by
conjugation or by transduction.
*R has two segments:
1) ‘RTF’ (the resistance transfer factor): that
controls the replication and
transferability of the plasmids.
2) ‘r’-determinant: that determines the resistance
to antimicrobials.
*(A bacterium having RTF+ but r- is meant that it is a
sensitive bacterium to antimicrobials)
DR. Mishuk Shaha

*It should be kept in mind that not all drug resistance is
attributable to genes carried by plasmids.
*Drug resistance may also arise from mutation in
chromosome which can be transferred by F-plasmids or
by a transducing phage.
DR. Mishuk Shaha

Transduction
*It is a phages/bacteriophages mediated genetic
recombination in bacteria.
*Phages/bacteriophages are viruses that parasitize
bacteria.
*In the simplest term, a transducing particle might be
regarded as bacterial DNA in a phage coat. Even a lytic
phage population may contain some particles in which
the phage coat surrounds DNA derived from the
bacterium.
*Such phage populations can be used to transfer DNA
from one bacterium to another.
DR. Mishuk Shaha

Lytic and lysogenic (temperate) cycles:
*Transduction happens through either the lytic cycle or the
lysogenic cycle.
* If the lysogenic cycle is adopted, the phage chromosome
is integrated (by covalent bonds) into the bacterial
chromosome, where it can remain dormant for thousands
of generations.
*If the lysogen is induced (by UV light for example), the
phage genome is excised from the bacterial chromosome
and initiates the lytic cycle, which culminates in lysis of
the cell and the release of phage particles.
*The lytic cycle leads to the production of new phage
particles which are released by lysis of the host.
DR. Mishuk Shaha

* Types of Transduction: 2 types
1. Generalized Transduction:
* Generalized transduction is the process by which any
bacterial gene may be transferred to another
bacterium via a bacteriophage, and typically carries
only bacterial DNA and no viral DNA.
* In essence, this is the packaging of bacterial DNA
into a viral envelope.
* This may occur in two main ways, recombination and
headful packaging.
DR. Mishuk Shaha

2. Specialized Transduction
* Specialized transduction is the process by which genes
that are near the bacteriophage genome may be
transferred to another bacterium via a bacteriophage.
* The genes that get transferred (donor genes) always
depend on where the phage genome is located on the
chromosome.
* This second type of recombination event which is the
result of mistakes in the transition from a virus'
lysogenic to lytic cycle is called specialized
transduction, and non-viral DNA is carried as an
insertion/substitution
DR. Mishuk Shaha

Recombinant DNA
technology/Genetic engineering
*Engineering is the application of science to social needs.
*Genetic engineering deals with the taking of genes from
one organism and planting them into another.
Basic steps in Genetic engineering:
*Selection and preparation of plasmids.
*Cutting of plasmid DNA by restriction endonuclease
restriction enzyme.
*Cutting of foreign DNA by restriction endonuclease
rstriction enzyme
DR. Mishuk Shaha

*Joining of the cut foreign DNA to the plasmid (cut) by the
enzyme ligase.
*Transformation of the recombinant plasmid into a
suitable bacterium.
*Recording growth of the transformed cell.
*Testing for the phenotypic expression of the desired
gene(s).
DR. Mishuk Shaha

Bacterial genetics

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