Bacterial Genomes

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• Bacterial Genomes
• Remember no nucleus!!
• Bacterial chromosome
• - Large ds circular DNA molecule
  haploid
• - E. coli has about 4,300 genes
  (4.2 Mb)

• 100x more DNA than the average virus
• 1000x less DNA than eukaryotic cell

- Chromosome is tightly coiled into dense
body nucleoid
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Prok Genome Size
Mega 106
Bacteria Size (Mbp) Escherichia
coli 4.64 Bacillus subtilis 4.20 Streptoco
ccus pyrogenes 1.85 Mycobacterium
genitalium 0.58
Archaea Size (Mbp) Methanococcus jannaschii
1.66 Sulfolobus solfactaricus 2.25 Pyrococcus
furiosus 1.75
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Euk Genome Size
Organism Mbp Homo sapiens 3,000 Drosophi
lia melanogaster 165 Plasmodium falciparum
23 Saccharomyces cerevisiae
12.07
Eukaryotes also have Mitochondrial
DNA Chloroplast DNA
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- Bacteria divide by simple division
binary fission - Division proceeded by
chromosome replication from single origin of
replication - E. coli cells can divide
every 20 min under optimal conditions - DNA
molecules are identical except for
mutations - Mutation rate 1 mutation/chromosome
/generation - With short generation time lots
of mutations 107-108 mutations/12 hours
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Extrachromosomal DNA - Many bacteria have
extrachromosomal molecules of DNA
plasmids - Plasmids contain an average of
10-50 genes - Cells can contain 1-100
plasmids
Resistance (R) plasmids - Usually carry
genes that detoxify antibiotics - Allows
bacteria to be resistant (R) to drugs
that would normally kill them - Also often
contain genes for sex pilus can be
transferred by conjugation (F plasmids)
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Recombination in Bacteria - Bacteria are
haploid, have only 1 copy of each gene
on circular chromosome - There are
mechanisms to introduce pieces of DNA
from one cell to another to produce a partial
diploid - Partial diploids, because
usually only small pieces of DNA with
only a few genes are transferred - The
foreign DNA in a partial diploid can replace
endogenous DNA in the chromosome by
homologous recombination
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Genetic recombination - exchange of genes between
two related chromosomes, forms new combinations
of genes Involves crossover event between
chromosomes
Results in hybrid chromosomes Each now has
properties of both original chromosomes In
eukaryotes, occurs during meiosis Increases
genetic diversity
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Can generate partial diploids in 3 different ways
Transformation - Bacteria take up naked
foreign DNA from the environment
- Consequences can be that mutant alleles are
replaced with wildtype alleles or vice
versa by homologous recombinationcrossi
ng over

• Not all bacteria can be naturally transformed
• Competence

• Can create competent cells in the lab

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Types of Transfer of Genetic Material
Genes can be passed from parental cell to
progeny cell vertical gene transfer Only
method of transfer in higher eukaryotes (yeasts
may be an exception) Also used by bacteria
Bacteria can also undergo horizontal gene
transfer Transfer of genetic material from one
cell to another Can result in a recombination
event Generates recombinant bacteria
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Transforming Principle Experiment 1928
Fred Griffith
Streptococcus pneumoniae
Smooth Virulent
Rough Avirulent
a-hemolysis of RBCs
No capsule!
Blood Agar Plate
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The Transforming Principle

• R cells were transformed
• Something in the S cells transformed the R cells
• The standard assumption was that proteins were
  responsible

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How were the Bacteria Transformed?
Rough colonies lacked functional gene for capsule
production DNA containing functional gene from
heat killed smooth bacteria taken up by rough
bacteria Recombination event replaced defective
capsule production gene with functional
gene Once rough bacteria can now make capsule
and are transformed to smooth colony virulent
phenotype
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Conjugation General features
- Transfer of genetic material between 2
bacteria that are temporally
joined - The donor cell transfer DNA
to the recipient cell - A sex pilus
from the male initially joins the 2
cells via cytoplasmic bridge -
Maleness is the ability to form a
sex pilus and donate DNA - Maleness
requires an F factor found either
on the bacterial chromosome or on
a plasmid
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Conjugation
Bacteria can exchange genetic information through
conjugation
Requires presence of fertility plasmid (F
plasmid) Contains genes required for production
of sex pilus

• Can connect two bacteria with pilus, one with
  plasmid (F) one without (F-)
• F plasmid transferred
• Converts F- to F

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How conjugation works - F factor is an
episome can exist as an autonomous
or integrated (into bacterial
chromosome) plasmid - The F factor
contains 25 genes mostly used to make
the sex pilus - Cells with the F factor
F conjugation donors - Cells without
the F factor F- conjugation recipients
- When F and F- meet, F donates the F factor
to F- cell and converts it to F
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F factor on plasmid - the plasmid is only
transferred during mating
F factor integrated into the bacteria chromosome
- occurs at a specific site - the
resulting cell is Hfr (High frequency of
recombination).
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Transduction - Occurs when phage picks up
piece of degraded bacterial chromosome
by mistake - The bacterial DNA is
transferred from one host to another by
the phage during infection
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More on phage during the virus lectures
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Regulation of Genes in Prokaryotes
In general, prokaryotic genes are organized (and
Expressed) as operons An operon consists of
Several genes that encode enzymes under the
control of a single promoter -
usually all enzymes needed for a specific
activity - all transcribed as one long
mRNA - polycistronic mRNA - mRNA
that contains that codes for more than one
gene within the same mRNA transcript

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promoter region
- site where RNA polymerase binds
- binding to promoter is necessary for
transcription of the mRNA that
encodes the enzymes
operator region -
binding site between the promoter and
first structural gene
- acts as an on-off switch
repressor protein - binds to
the operator region - prevents
transcription inducer molecule
- binds to repressor allows
transcription
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Transcriptional Control in Prokaryotes

• Lac Operon
• Escherichia coli

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Genes in the lac operon are designed to breakdown
and import lactose
Disaccharide
b galactosidase
Monosaccharide
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Three lactose metabolizing enzymes are under the
control of one promoter
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No transcription
Repressor protein binds to operator
Repressor is produced constitutively
No need to make b-gal other enzymes when
lactose is not present
Negative Control
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allolactose
A small amount is converted to allolactose
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Positive Control of the Lac Operon

• - Activator protein
• CRP cAMP Receptor Protein
• Concentration of glucose is low
• cAMP accumulates
• CRP/cAMP binds to the promoter
• Theres a special sequence / binding location
• Maximal rates of transcription occur
• Synthesize a lot of b-gal other enzymes

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Positive Control High Glucose

• - There is little cAMP
• - CRP can not be activated
• - E. coli prefers glucose
• If theres plenty of glucose
• No reason for produce b-gal
• The lac operon is shut down in the presence of
  glucose

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glucose ? cAMP ? Reduced transcription