BACTERIAL GENETICS

BY A. ARUN SUBRAMANIAN
1ST YEAR MICROBIOLOGY
 OBJECTIVES
At the end of the session, you should be able to answer the following
questions
. What is genotype and phenotype?
. Mutation and its types
. Conjugation, Mechanism of Conjugation
. Transformation principle and Griffith experiment analysis
. Transduction, types of transduction
 GENOTYPE PHENOTYPE
It is the genetic makeup of an organism It is the morphology, biochemical and
physiological properties, behavior and products
of behavior of an organism
Genotype can be determined by observing Phenotype can be determined by observing
DNA by genotyping methods outward characters.

Genotype completely depends on the gene Phenotype depends on the genotype and
sequences. environmental factors

It is inherited by the offspring. It is not inherited by the offspring. But natural

selection can alter the genetic makeup.
It consists of all hereditary information that is It only consists of expressed genes.
the expressed and suppressed genes.
Genotype :

* Genotype is the complete set of genes carried by a particular

organism.
* Thus, it is a part of the sequences of the genome or the genetic
makeup of that organism.
* The genotype carries the instructions for development and
functioning of a cell.
* Hence, it is referred to as the “blueprint” of a cell.
* These instructions are written by the aid of the genetic code.
• Genotype is one factor that determines the characteristics,
appearance and behavior of a particular organism.
• The appearance and the behavior can be altered by inherited
epigenetic factors and environmental factors.
• Therefore, two individuals carrying similar genotypes can be different
in observable characteristics.
• However, the genotype is inherited through the progeny via
reproduction.
• A genotypic assay may include PCR, RFLP, nucleic acid hybridization,
DNA sequencing or DNA microarray techniques
• Phenotype :

• Phenotype is the physical manifestation of an organism, including the observable

structure, function and the behavior.
• The genotype of an organism determines its molecules, macromolecules, cells,
metabolism, energy utilization, organs, tissues, reflexes and the behavior.
• The genotype, together with other two factors: epigenetic and environmental factors,
determines the phenotype of that organism.
• Simply, the phenotype is what you see or the observable expression of genes
combining with environmental influence.
• The occurrence of more than one phenotypes for a particular morphological trait is
referred to as phenotypic polymorphism.
 MUTATION
• Mutations are heritable changes in the genetic coding instructions of
DNA. They are essential to the study of genetics and are useful in
many other biological fields.
• Mutations can be beneficial or harmful to the cell.
• The changes in the cell may be due to
• Substitution of a nucleotide
• Deletion of a nucleotide
• Addition of a nucleotide
• Spontaneous mutations: As a result of errors in DNA replication

• Induced mutations: By exposure to radiation, chemicals, viruses, or

other mutagenic agents
TYPES OF MUTATION
Missense Mistake in the DNA code, one of the DNA base pairs is changed

Nonsense Single change in DNA code produces stop codon, prematurely terminates protein synthesis

Insertion Addition of one (or more) nucleotide base pairs into the DNA sequence

Deletion A piece of DNA is removed from the sequence

Frameshift Addition or deletion mutation results in a change to a gene's reading frame

Duplication DNA is abnormally copied

• MISSENSE
MUTATION: A genetic
alteration in which a
single base pair
substitution alters
the genetic code in a
way that produces
an amino acid that is
different from the
usual amino acid at
that position.
• Like a missense mutation, a
nonsense mutation also
involves a single alteration to
the DNA base pair. However,
in the case of a nonsense
mutation, this single change
results in the production of a
stop codon, thereby
terminating protein synthesis
prematurely. The result A
shortened protein that may
function, but also may not.
• Insertion are mutations in
which extra base pairs are inserted into a
new place in the DNA.
• Eg: Huntington's disease and the fragile
X syndrome are examples of insertion
mutation wherein trinucleotide repeats
are inserted into the DNA sequence
leading to these diseases.
• Deletions are
mutations in which a
section of DNA is lost or
deleted.
• Eg: 22q11.2 deletion
syndrome is caused by
the deletion of some
bases of chromosome
22. This disease is
characterized by cleft
palate, heart defects,
autoimmune disorders
etc.
• A frameshift
mutation is a
genetic mutation
caused by a
deletion or insertion
in a DNA sequence
that shifts the way
the sequence is
read.
• In the case of a
duplication
mutation, a piece of
DNA is (abnormally)
copied. This might
happen once or several
times, and can
therefore impact the
functionality of the
encoded protein.
 GENE TRANSFER
• Genetic information can be transferred from one bacterium to
another by several methods
• 1. Horizontal gene transfer
TRANSFORMATION
TRANSDUCTION
CONJUGATION

2. Vertical gene transfer

 RECOMBINATION
 CONJUGATION

• Conjugation involves the transfer of DNA from donor bacterium to

recipient bacterium by cell contact.
• The first step in conjugation involves a conjugative pilus (sex pilus or F
pilus) on the donor bacterium (called F+) binding a recipient
bacterium lacking a conjugative pilus (called F−)
1.Plasmids are small, circular pieces of DNA that are separate
and replicate independently from the bacterial chromosome.
2.Plasmid carries genes associated with specialized functions
such as drug resistance.
3.Plasmids may encode genes that mediate their transfer from
one organism to another but not all plasmids are capable of
conjugative transfer
4.Plasmids that can transmit from one cell to another cell
independently are called self-transmissible plasmids. Non-
transmissible plasmids are not able to go through independent
transmission.
• The F plasmid also called the ‘fertility factor’ confers donor
characteristics (sex pilus) to bacterial cells.
1.F+ strains: Bacteria that have F plasmid are referred to as F+
or male/donor.
2.F- strains: Those that do not have F plasmid are called F- or
female/recipient.
3.Hfr strains: Arise from F+ strains when the F plasmid is
incorporated into the bacterial chromosomes at one of several
possible sites.
4.F’ (prime) strains: F plasmid with a fragment of chromosomes
with it.
 MECHANISM OF CONJUGATION

• The plasmid DNA is nicked at a specific site called the origin of transfer.

• Pair formation - The tip of the sex pilus encounters the recipient, and a
conjugation bridge is formed between the two cells, through which the
plasmid DNA will pass from the donor to the recipient

* A single strand of plasmid DNA passes through the conjugation bridge and
enters the recipient where the complimentary strand is synthesized by a rolling
circle mechanism
This results in the transfer of a F+ plasmid (coding only for a sex pilus),
but not chromosomal DNA, from a male (F+) donor bacterium to a
female recipient (F-) bacterium.

The recipient F- then becomes a F+ (male) and can make a sex pilus.

Other genes present on the plasmid, such as those coding for antibiotic
resistance, may also be transferred during this process.
 TRANSFORMATION
• Transformation is a method of genetic recombination
in which a naked DNA from a donor bacteria is
transferred to a competent recipient bacteria and
incorporated into chromosome of the latter.
• e.g. in Bacillus, Haemophiles, Neisseria, Pneumococcus.
• Transformation occurs in nature.
• It is widely used in recombinant DNA technology.
• In Gram+ve bacteria the DNA is taken up as a single stranded
molecule and the complementary strand is synthesized in the
recipient.
• In Gram-ve bacteria double stranded DNA is transformed
 THE GRIFFITH
EXPERIMENT
• Conducted in 1928 by Frederick Griffith; was the first experiment
suggesting that bacteria can transfer genetic information through a
process known as transformation

• Griffith was studying the possibility of creating a vaccine for

pneumonia.
• He used two strains of pneumococcus (Streptococcus pneumoniae)
bacteria which infect mice – a type III-S (smooth) which was virulent
(causing disease), and a type II-R (rough) strain which was
nonvirulent.
• The III-S strain synthesized a polysaccharide capsule that protected
itself from the host's immune system, resulting in the death of the
host, while the II-R strain did not have that protective capsule and
was defeated by the host's immune system
• In this experiment, bacteria from the type III-S strain were killed by
heat, and their remains were added to type II-R strain bacteria, While
neither alone harmed the mice, the combination was able to kill its
host.
• Griffith was also able to isolate both live II-R and live III-S strains of
pneumococcus from the blood of these dead mice.
• Griffith concluded that the type II-R had been "transformed" into the
lethal III-S strain by a "transforming principle" that was somehow
part of the dead III-S strain bacteria.
• He had no idea of that transforming agent
• Today, we know that the "transforming principle" Griffith observed
was the DNA of the III-s strain bacteria.
• While the bacteria had been killed, the DNA had survived the heating
process and was taken up by the II-R strain bacteria.
• The III-S strain DNA contains the genes that form the smooth
protective polysaccharide capsule.
• Equipped with this gene, the former II-R strain bacteria were now
protected from the host's immune system and could kill the host.
 TRANSDUCTION
• : Gene transfer from a donor to a recipient bacteria through a
bacteriophage

• Types of transduction
• * Generalized
• * Specialized
 GENERALIZED
TRANSDUCTION

• Generalized transduction can occur by both lytic or

lysogenic cycle. Here, any random part of DNA gets
packed in bacteriophages by mistake along with the
viral genome.

• It occurs at the lytic stage of the phage life cycle.

• When the virus-containing bacterial DNA infects another
cell, it can get inserted into the host genome or if it was
a plasmid, then it can reform the plasmid.

• Generalized transduction is used to study linkage

information, gene mapping, comparing genomes of two
different bacteria, mutagenesis, etc.

• Example of generalized transduction

includes E.coli transduction by P1 phage.
 RESTRICTED
TRANSDUCTION
• Specialized transduction can occur only through the
lysogenic cycle, i.e., by temperate phage.
• Here, only the specific part of the bacterial DNA is
packed into the virus.
• It occurs when the prophage, i.e., viral DNA, which gets
inserted into the bacterial genome in the lysogenic
cycle excises.
• When prophage excises from bacterial DNA, some parts
of bacterial DNA, which are flanked on both sides of the
prophage are also excised.
• Here, the newly packed phage genome consists of both
bacterial and viral genome.
• Later, when the virus with the recombinant genome
infects a new bacterial cell, the bacterial gene also gets
inserted into the host genome with the viral genome
through lysogeny.
• The recipient cell now shows the newly acquired
characteristics.
• Specialized transduction is commonly used for isolation
and insertion of genes of choice.

includes E.coli transduction by 𝝀 phage.

• Example of specialized transduction
• SUMMARY
The genotype is a set of genes in DNA responsible for unique
trait or characteristics
Phenotypes are responsible for the organism’s
observable characteristics.
A key difference between phenotype and genotype is
that, while genotype is inherited from the parents, but
the phenotype is not.
Mutations is heritable change in DNA caused by change
in nucleotide sequence
Conjugation the process whereby a donor male
bacterium conjugates or makes physical contact via sex
pilus with the recipient female bacterium and transfers
the genetic elements into it.
 Transformation is the transfer of genetic information
through free DNA
Transformation experiment was performed by Griffith in
1928 with
S. pneumoniae with its two strains the type II non
capsulated (R)and
type I capsulated(S)
Transduction is the phenomenon of transfer of a
portion of DNA from one bacterium to another by
bacteriophage.
Two types of transduction are
Generalized – involves any segment of donor DNA randomly
Restricted – when specific phage transduces only a particular genetic
trait.