Transcript Recomn in Bacteria and Viruses

30. Genetics and recombination in
bacteria
Lecture Outline 11/16/05
• Replication in bacteria
• Types of recombination in bacteria
– Transduction by phage
– Conjugation (“mating”)
• F+ plasmids
• Hfr strains
– Transformation of raw DNA
• Evidence for recombination in nature
– Resistance plasmids
The Bacterial Genome and Its
Replication
• The bacterial chromosome
– Is usually a circular DNA molecule with few
associated proteins
• In addition to the chromosome
– Many bacteria have plasmids, smaller circular DNA
molecules that can replicate independently of the
bacterial chromosome
Genetics fo Bacteria
• Use huge numbers of
individuals (billions) To find
very rare events
• Few morphological traits
– Antibiotic resistance
– “Auxotrophs” cannot synthesize
essential nutrients (arg- or trp-)
– “Prototrophs” have normal
synthesis (arg+, trp+)
Replication of the circular chromosome
Origin of
replication
Replication always
starts at a certain place
Replication
fork
Normal replication
fork for DNA
synthesis
Termination
of replication
Figure 18.14
Bacterial cells usually divide asexually
by binary fission
but they can occasionally exchange genes:
EXPERIMENT
Mix two mutant strains: Arg+ Trp- and Arg- Trp+.
Grown them on complete media. After a short while, test them
on culture medium without Trp or Arg.
Mixture
Mutant
strain
arg+ trp–
Figure 18.15
Mutant
strain
arg trp+
Now test them on minimal culture medium
RESULTS
Mixture
Why do they need
the control plates?
Mutant
strain
arg+ trp–
Mutant
strain
arg– trp+
No
colonies
(control)
Colonies
grew
No
colonies
(control)
CONCLUSION
To grow on minimal medium, the cell must be able to
make both Arginine and Tryptophan (Arg+, Trp+).
--> Evidence for genetic transfer of one of those genes to
the other strain.
Four ways bacteria can exchange genes
1. Transduction
Phage can transfer
bacterial genes
between cells
1. Phage virus infects
A+B+ cell
2. Reproduction
and lysis
Once in a while host
DNA is mistakenly
packaged in a capsid
3. Transfer of a+ DNA from phage to new cell
2. Conjugation
– direct transfer of genetic material between bacterial
cells that are temporarily joined
Donor cell
contains F+
plasmid
Recipient cell
is F(has no plasmid)
One way transfer
Figure 18.17
Sex pilus
1 m
Conjugation and transfer of an F plasmid
from an F+ donor to an F recipient
F Plasmid
Bacterial chromosomes
F+ cell
F+ cell
Mating
bridge
F+ cell
F– cell
F+ cell can form a
mating bridge with an F– cell
and transfer its F plasmid.
Figure 18.18a
Single strand of the
F plasmid breaks at a
specific point and
moves into the recipient cell.
Both
cells are now F+.
Donor F+ cell
Synthesis of
complementary
strand in recipient
Structure of F plasmid
These genes play a role in the transfer of DNA
They are thus designated tra and trb followed by a capital letter
Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display
3. High Frequency Recombination
(Hfr cells)
F factor can sometimes become integrated in to a
bacterial chromosome.
F+
Cell is F+ because it
has all of the F factor
genes
MUCH more likely to transfer chromosomal
genes to F- cell during conjugation
Conjugation of
Hfr cell with
F- cell)
See this in action
Usually carries some chromosomal DNA along with it
when it is transferred to an F– cell
Conjugation and transfer of part of the bacterial
chromosome from an Hfr donor to an F– recipient
Hfr cell
F+ cell
F factor
1
Hfr cell
A+
The circular F plasmid in an F + cell
can be integrated into the circular
chromosome by a single crossover
event (dotted line).
B+
C+
The resulting cell is called an Hfr cell
(for High frequency of recombination).
D+
C+
B+
D+
2
D+ C+
A+
B+
A+
D+ C+
B+
B+
A+
B–
A+
A+
F– cell
3
B–
C–
A–
B+
D–
Since an Hfr cell has all
the F-factor genes, it can
form a mating bridge with
an F– cell and transfer DNA.
B–
Figure 18.18b
A–
B+
A+
Two crossovers can result
in the exchange of similar
(homologous) genes between
the transferred chromosome fragment
(brown) and the recipient cell’s
chromosome (green).
B–
C–
A–
A+
D–
4 A single strand of the F factor
breaks and begins to move
through the bridge. DNA
replication occurs in both donor
and recipient cells, resulting in
double-stranded DNA
Temporary
partial
diploid
7
C–
B–
C– D–
A–
5 The location and orientation
of the F factor in the donor
chromosome determine
the sequence of gene transfer
during conjugation. In this
example, the transfer sequence
for four genes is A-B-C-D.
B–
D–
A+
B+
C–
A–
D–
6
C–
A–
D–
The mating bridge
usually breaks well
before the entire
chromosome and
the rest of the
F factor are transferred.
Recombinant F–
bacterium
8 The piece of DNA ending up outside the
bacterial chromosome will eventually be
degraded by the cell’s enzymes. The recipient
cell now contains a new combination of genes
but no F factor; it is a recombinant F – cell.
Integration of F+ plasmid into a
chromosome
Transformation
• Transformation
– uptake of naked, foreign DNA from the
surrounding environment
• Remember Griffith’s experiment with heat killed
bacteria and mice?
Does this happen in nature?
• In E. coli and Salmonella, roughly 17% of
their genes have been acquired from other
species (over 100 million years . . . )
• Such “horizontal transfer” is an important
issue for the spread of antibiotic resistance
Spread of Atrizine decomposing
bacteria
• A few bacterial species are capable of
metabolizing the synthetic herbicide
Atrizine
• All have nearly identical genes.
Atrizine catabolism plasmid
Genes DEF in an operon
Dispersed atrizine
catabolism genes
(ABC) acquired
separately?
Transposons
flank these
genes
Resistance mechanisms
Antibiotic
Method of resistance
-----------------------------------------------------------------------Chloramphenicol
Tetracycline
B-lactams, Erythromycin,
B-lactams, Erythromycin
Aminoglycosides, Chloramphenicol,
B-lactams, Fusidic Acid
Sulfonamides, Trimethoprim
Sulfonamides, Trimethoprim
Bleomycin
reduced uptake into cell
active efflux from the cell
eliminates or reduces binding of antibiotic to target
hydrolysis
inactivation of antibiotic by enzymatic modification
sequestering of the antibiotic by protein binding
metabolic bypass of inhibited reaction
overproduction of antibiotic target (titration)
binding of specific immunity protein to antibiotic
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