Download Genetic Exchange

Genetic exchange
• Mutations
• Genetic exchange: three mechanisms
• Transposons
Mutations and Adaptation
• In the course of DNA replication mutations can arise in the bacterial
genome.
• These can be point mutations in which one base is substituted by
another. Point mutations in a coding sequence can lead to, no
change in the protein (silent mutation), a change in an amino acid or
the conversion of an amino acid coden to a STOP codon.
• Deletion and insertion and insertion mutations normally have a
deleterious effect.
• In summary point mutations can only be used to ”tinker” with existing
genes and are not a viable mechanism for the aquisition of new
genetic properties.
• In a given population of bacteria there will always be a certain level
of mutants and these will only dominate if they can grow more
quickly than the wild type.
Genetic Exchange
• There are three different natural processes by which bacteria can
gain new genetic material (DNA).
• Transformation in which DNA is taken up from the environment
• Conjugation in which a plasmid is transferred from one bacteria to
another.
• Transduction in which the transfer of DNA from one bacteria to
another is mediated by a bacteriophage.
Demonstration of transformation
Transformation of avirulent Streptococcus pneumoniae to a virulent type.
Avery, MacLeod and McCarty 1944
Demonstration of transformation
Demonstration
that the
transforming
factor is DNA.
Natural transformation competence
• Many different bacteria are naturally competent. Some are
competent all the time, others only at specific stages in the
bacteria’s growth phase.
• Examples are, Streptococcus pneumonia, Bacillus subtilis,
Hemophilus influensa, Neisseria gonorrhoeae.
• Many other bacteria have been shown to contain the genes for
natural competence but have never been observed to do so.
• The molecular mechanism of transformation has been studied in
some detail in a few species. The subtrate is double stranded linear
DNA but only one of the DNA strands enters the cell. Recombination
of this single stranded DNA into the bacterial chromosome is
necessary for expression. Normally only DNA from a closely related
species can be taken up by transformation and integrated into the
chromosome.
Details of homologous recombination
What are the consequences of
transformation
Existing genes can be extensively modified by
exploiting existing variation within a population.
It has been shown that this can be important for
proteins associated with host interactions in
pathogenic bacteria.
Target genes for antibiotics can be quickly
modified and antibiotic resistence can be
established.
Artificial competence
• All bacteria can be treated (chemically / electroporation) such that
they become competent and can take up DNA.
• The substrate here is normally a plasmid which can replicate in the
bacteria cytoplasm. This is one of the corner stones of recombinant
DNA technology.
• If linear DNA is artificially transformed into a bacteria then it must be
incorporated into the bacterial chromosome by double
recombination in order to be expressed.
• Note that we are now speaking about double stranded DNA.
Types of homologous recombination in
bacteria
Double recombination
Introduction of a mutation into a bacterial chromosome from a
piece of DNA acquired by transformation. This is the basis for gene
knock out.
Gene replacement
Introduction of a new gene into a bacterial chromosome by
transformation. General recombination occurs between homologous
sequences that flank the gene.
Plasmids in general
• Some plasmids cannot be transferred by
conjugation.
• Some plasmids cannot be transferred by
conjugation but they can be helped to transfer if
a conjugative plasmid is also present in the cell.
• Some plasmids contain all the necesary genes
to mediate their transfer to another bacteria by
conjugation.
• Some conjugative plasmids have a narrow host
range , others are broad range.
Plasmids of pathogenic bacteria
Genome maps of some
plasmids found in pathogenic
bacteria. (A) A widely
distributed R plasmid, RK2. (B)
Plasmid pCG86 of pathogenic
E. coli. The gene product or
function of the genes is
indicated. β-Lactam antibiotics
include ampicillin.
Ti plasmid
Genome map of the
Ti plasmid of
Agrobacterium
tumefaciens,
showing the gene
product or function of
the genes.
Phytohormones are
responsible for the
induction of plant
tumors.
Genetic Exchange
• There are three different natural processes by which bacteria can
gain new genetic material (DNA).
• Transformation in which DNA is taken up from the environment
• Conjugation in which a plasmid is transferred from one bacteria
to another.
• Transduction in which the transfer of DNA from one bacteria to
another is mediated by a bacteriophage.
Bacterial conjugation
Transfer of F plasmid from
donor to recipient cells by
conjugation. Once transfer is
complete, both cells have an
intact copy of F plasmid and
can act as donors.
The F plasmid is large, ~100
kb and contains about 100
genes.
High-frequency recombinant cells
High-frequency recombinant cells
Transfer of chromosomal genes
into a recipient bacterium.
Orientation of the inserted F
plasmid in the opposite direction
from that shown here would
allow early transfer of genes e,
b, and c and later transfer of d
and a. Relative locations of
genes in the bacterial
chromosome can be mapped by
mixing donor and recipient cells,
interrupting the mating at
various times, growing the cells
on appropriate media, and
identifying the transferred
genes.
F' cells
Formation of an F' cell from
an Hfr cell, and transfer of a
bacterial chromosome
segment to a recipient cell.
Consequences of conjugation
A bacteria cell can get many new genes and in turn new
genetic properties when it gets a new plasmid.
In some cases these can be incorporated into the genome
by recombination and they thus become part of the
genome (plasmids can be lost).
Plasmids play an important roll in the transfer of antibiotic
resistence between bacteria. A deadly combination if
they are pathogenic.
Bacteriophage
Lets take a general look at bacteriophages before
we look at the roll of bacteriophages in genetic
exchange.
Complex virus
Graphic representation of T4 virus (phage).
Viral reproduction: the lytic cycle
Generalized schematic for
viral reproduction in a host
bacterium, through the lytic
cycle.
In the lytic cycle, the virus
(phage) multiplies in the
host cell and the progeny
viruses are released by
lysis of cell.
Viral reproduction: the lysogenic cycle
Generalized schematic for
viral reproduction in a host
bacterium, through the
lysogenic cycle.
In the lysogenic cycle, viral
DNA is integrated into the
host genome and replicates
as the chromosome
replicates, producing
lysogenic progeny cells
Genetic Exchange
• There are three different natural processes by which bacteria can
gain new genetic material (DNA).
• Transformation in which DNA is taken up from the environment
• Conjugation in which a plasmid is transferred from one bacteria to
another.
• Transduction in which the transfer of DNA from one bacteria to
another is mediated by a bacteriophage.
Generalized transduction: Lytic phage
Specialized transduction: Lysogenic phage
Consequences of transduction
Specialized transduction can only transfer genes that flank
the specific insertion site and as such do not contribute
many new genes to the bacteria.
Generalized transduction can be instrumental in the
transfer of 50-100 new genes and make dramatic
changes to the properties of the bacteria. Transduction
plays an important roll in the transfer of, antibiotic
resistence and pathogenicity factors. This can be a
deadly combination.
Insertion elements and transposons
Insertion sequences (IS) are short DNA sequences, about
700 to 5000 bp which can move from one location in a
DNA sequence to another. They have short 16-41 bp
inverted repeats on their ends. They encode a
transposase which catalyses site-specific recombination.
Simple transposons are mobile genetic elements in which
a one or more genes are flanked by two insertion
sequences.
Composite transposons
Structures of some bacterial transposable elements.
(A) A composite transposon contains antibiotic genes flanked by two
insertion sequences as direct or inverted repeats Shown here is the
Tn5 transposon, with inverted repeats.
(B) The Tn3 transposon.