Genetic exchange

1
Genetic exchange

• Mutations
• Genetic exchange three mechanisms
• Transposons

2
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.

3
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.

4
Demonstration of transformation
Transformation of avirulent Streptococcus
pneumoniae to a virulent type. Avery, MacLeod and
McCarty 1944
5
Demonstration of transformation
Demonstration that the transforming factor is DNA.
6
Natural transformation competence

• Many different bacteria are naturally competent.
  Some are competent all the time, others only at
  specific stages in the bacterias 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.

7
Details of homologous recombination
8
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.

9
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.

10
Types of homologous recombination in bacteria
11
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.
12
Gene replacement
Introduction of a new gene into a bacterial
chromosome by transformation. General
recombination occurs between homologous sequences
that flank the gene.
13
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.

14
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. ß-Lactam antibiotics include
ampicillin.
15
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.
16
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.

17
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.
18
High-frequency recombinant cells
19
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.
20
F' cells
Formation of an F' cell from an Hfr cell, and
transfer of a bacterial chromosome segment to a
recipient cell.
21
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.

22
Bacteriophage

• Lets take a general look at bacteriophages before
  we look at the roll of bacteriophages in genetic
  exchange.

23
Complex virusGraphic representation of T4 virus
(phage).
24
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.
25
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
26
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.

27
Generalized transduction Lytic phage
28
Specialized transduction Lysogenic phage
29
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.

30
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.

31
Composite transposons

• Structures of some bacterial transposable
  elements.
• 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.
• The Tn3 transposon.