Transcript first of four for Chapter 9

Chapter 9
Genetics of
Bacteria and
Their Viruses
Jones and Bartlett Publishers © 2005
Mobile DNA
• Plasmids
• Insertion sequences and transposons
• Integrons
• R plasmids
Mobile DNA
• Plasmids – can be circular or linear.
• Not essential for bacterial cells
• Can be 1-2 copies or up to 50 copies per host
cell.
• Range in size from a few kb to a few
hundred kb.
• Plasmids depend on host DNA replication
enzymes.
F plasmid: Conjugation
• In E. coli, a large plasmid can be
transferred between cells (distinct from
fission and replication).
• F factor (F for fertility)
• Low copy number, 100 kb
• F+ vs F• Pilus (pl., pili), a tube-like structure that
connects two cells undergoing conjugation.
• 20 genes necessary for pilus assembly and
DNA transfer.
Conjugation in E. coli between a male (F+)
and a female (F-) cell
Insertion sequences and transposons
• Transposable elements are responsible for
gene mobilization. How they work to move
genes, and act as mutagens will be discussed
later.
• In bacteria, the smallest transposable
elements are insertion sequences, or IS
elements.
• IS elements are 1-3 kb and encode a
transposase protein and a few related
proteins.
Tn5 (a composite transposon) has IS elements at each
end (IS50 left and IS50 right) and drug resistance
genes in the middle
IS elements are the simplest known fully functional transposons
(they have one gene flanked by inverted repeats). Neo, ble and str
refer to the antibiotics neomycin, bleomycin and streptomycin.
Mobilization of non-conjugative plasmids
• Transposons can move between host DNA of
a bacterium and plasmids in the cell.
• E. coli contains 1-6 genomic copies of each
of 6 naturally occurring IS elements.
• In cells with a particular IS element, 2060% contain the IS element in plasmids.
• These IS elements in plasmids can serve as
recombination sites (homology).
Recombination between 2 transposons leads to the
fusion of 2 plasmid circles creating a larger circle
Cointegrate = a composite plasmid.
In this way, nonconjugative plasmids can
catch a ride with a conjugative plasmid.
Site-specific recombination involves 2 short target
sequences and an enzyme capable of recombining them
Cre recombinase
recognizes the loxP
site.
Site-specific recombination and
antibiotic resistance
• Multiple-antibiotic resistance units can be
assembled to form integron.
• An integron is a DNA element which
encodes a site-specific recombinase as well
as a recognition sequence that allows other
similar elements to join the integron.
• A cassette is a circular antibiotic-resistancecoding region flanked by a recognition
region.
One or more short circular DNA molecules may be
inserted in a DNA target by site-specific recombination
R plasmids
• Some conjugative plasmids in nature have
accumulated different transposons
containing multiple antibiotic-resistance
genes (or transposons with integrons with
the resistance genes).
• These plasmids with multiple resistance
genes are called R plasmids, and cause
clinical trouble.
Bacterial Traits
• Bacteria are small: about 1/1000th the
volume of eukaryotic cells.
• Escherichia coli are about 1 mm long by 0.5
mm in diameter.
• With favorable temperature and nutrients,
bacteria can divide every 20-60 minutes.
• They are haploid, with only one allele at
each locus.
• They make clones in culture.
Bacterial Traits
• E. coli can grow either on solid media on a
Petri plate, or in liquid culture.
• A single cell of E. coli that doubles every
hour can make over 10 million cells in 24
hours.
• The colony will be visible on a plate, and
will be a clone
Bacterial genetics
• Mutants of bacteria exist:
– Unable to use certain nutrients
– Auxotrophic strains (wild type are
prototrophs and can grow on a
minimal medium)
– Sensitivity or resistance to drugs or
phage.
– Conditional mutants (temperature
sensitivity).
– Carbon source mutants
Types of genetic transfer in bacteria
• Transformation – does not require contact;
one cell lyses, another takes up the DNA
from the lysed cell.
• Conjugation – requires actual physical
contact between the donor cell and the
recipient.
• Transduction – involves a bacteriophage.
Genetic transformation can be used to
create genetic maps in bacteria