Click to add Text Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Brock Biology of Microorganis ms Twelfth Edition Madigan / Martinko Dunlap / Clark Principles of Bacterial Genetics Chapter 11 Professor Bharat Patel
Click to add TextCopyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Brock Biology of Microorganisms
Twelfth Edition
Madigan / Martinko
Dunlap / Clark
Principles of Bacterial Genetics
Cha
pter
11
Professor Bharat Patel
Click to add TextCopyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Brock Biology of Microorganisms
Twelfth Edition
Madigan / Martinko
Dunlap / Clark
Principles of Bacterial Genetics
Cha
pter
10
Professor Bharat Patel
Thirteenth Edition
Madigan / Martinko
Stahl / Clark
Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings
NOTE
1. The following is a summary and are not full notes for the Lecture on “Principles of Genetics”. This summary is a study guide only and it is therefore recommended that students attend and take notes during the lectures.
2. There are differences in the content of the chapters of the two different editions of the recommended text book
3. The lecture & summary may not follow the same content as is in the book chapter
4. There is extra content that has been sourced from other resources
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CONTENT
The lecture content is divided into 3 parts:I. Bacterial Chromosomes & Plasmids
• Physical location of the genes II. Mutation
• Alterations in the genetic material Chemical, Physical
III. Genetic Transfer• Gene transfer & exchange mechanisms
Conjugation Transduction Transformation
• Gene exchange mechanisms
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Note:
• Most of the techniques described here were used between 1950-80, but advances in the past three decades in cloning and sequencing has revolutionised studies on genomes & gene organisation:
• Developments in molecular biology: Manual sequencing & Automated 1st generation sequencers
1970 – 2008: $1-2 million per microbial genome2nd generation sequencers (current)
Since 2009: $5,000 per microbial genome3rd generation sequencers
early next year, semi-conductor real-time technology $1,000 per human genome
• Genomes OnLine Database (GOLD)- http://genomesonline.org – lists all genome sequencing projects.
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I. Genetics of Bacteria and Archaea
Lecture Content
11.1 Genetic Map of the Escherichia coli
Chromosome
11.2 Plasmids: General Principles
11.3 Types of Plasmids and Their Biological
Significance
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11.1 Genetic Map of the Escherichia coli Chromosome
Escherichia coli a model organism for the study of biochemistry, genetics, and bacterial physiology
The E. coli chromosome (strain MG1655, derivative of K-12) was been mapped using Conjugation (initial mapping) Transduction (phage P1) Molecular cloning & sequencing Next Generation Sequencing (NGS) (most recent)
E. coli is (gram -ve) is inefficient at transformation unlike Bacillus (gram +ve)
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Circular Linkage Map of the Chromosome of E. coli K-12
Figure 11.1
Original map used distance (centisomes)
0 – 100 mins, 0 = arbitrary & set at thrABC (based on transfer by conjugation)
Also shows kilobase pairs (kb) from sequencing studies
Replication starts at oriC (84min)
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11.1 Genetic Map of the Escherichia coli Chromosome
Some Features of the E. coli Chromosome Many genes encoding enzymes of a single biochemical
pathway are clustered into operons
Operons are equally distributed on both strands
Transcription can occur clockwise or anticlockwise
~ 5 Mbp in size
~ 40% of predicted proteins are of unknown function
Average protein size is ~ 300 amino acids
Insertion sequences (IS elements) are present
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Genomes of pathogenic E. coli contain PAIs.
Fig13.13
Genome size is indicated in the centre. The outer ring shows gene by gene comparison with all 3 strains: common genes (green), genes in pathogens only (red), genes only in 536 (blue)
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Pan Genome Versus Core Genome
Figure 13.14
Core genome is in black & is present in all strains of the same species.
The pan genome includes elements (genes) that are present in one or more strains but not in all strains.
one coloured wedge = single insertion
two coloured wedges = alternative insertions possible at the site but only can be present
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Plasmids
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11.2 Plasmids: General Principles
Plasmids: Genetic elements that replicate independently of the host chromosome
Small circular or linear DNA molecules
Range in size from 1 kbp to > 1 Mbp; typically less than 5% of the size of the chromosome
Carry a variety of nonessential, but often very helpful, genes
Abundance (copy number) is variable
Plasmid
Plasmid
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11.2 Plasmids: General Principles
A cell can contain more than one plasmid, but it cannot be closely related genetically due to plasmid incompatibility
Many Incompatibility (Inc) groups recognized Plasmids belonging to same Inc group exclude each
other from replicating in the same cell but can coexist with plasmids from other groups
Borrellia burgdorferi (causes Lyme disease) - 17 different circular & liner plasmids
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11.2 Plasmids: General Principles
Some plasmids (episomes) can integrate into the cell chromosome; similar to prophage integration – replication is under the control of the host cell
Host cells can be cured of plasmids by agents that interfere with plasmid (but not cell) replication
Acridine orange or can be spontaneous
Conjugative plasmids can be transferred between suitable organisms via cell-to-cell contact
Conjugal transfer controlled by tra genes on plasmid
Plasmid replicate up to 10 times faster than host cell DNA due to their small size
unidirectional (one fork) or bi-directional (two forks)
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11.3 Types of Plasmids and Their Biological Significance
Genetic information encoded on plasmids is not essential for cell function under all conditions but may confer a selective growth advantage under certain conditions
Plasmids are transferred by conjugation (refer to Conjugation later) – provide cells with additional “coping and fighting” strategies
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Examples of Phenotypes Conferred by Plasmids
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Examples of Phenotypes Conferred by Plasmids
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11.3 Types of Plasmids and Their Biological Significance
R plasmids Resistance plasmids; confer
resistance to antibiotics and other growth inhibitors
Widespread and well-studied group of plasmids
Many are conjugative
Outer ring: resistance genes (str streptomycin, tet tetracylcine, sul sulfonamides, & other genes (tra transfer functions, IS insertion sequence, Tn10 transposon). Inner ring: Plasmid size = 94.3 kb
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11.3 Types of Plasmids and Their Biological Significance
In several pathogenic bacteria, virulence characteristics
are encoded by plasmid genes
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11.3 Types of Plasmids and Their Biological Significance
Bacteriocins
Proteins produced by bacteria that inhibit or kill closely
related species or even different strains of the same species
Genes encoding bacteriocins are often carried on plasmids
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11.3 Types of Plasmids and Their Biological Significance
Plasmids have been widely exploited in genetic engineering for biotechnology
Plasmids are transferred by conjugation (refer to Conjugation later) – provide cells with additional “coping and fighting” strategies
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Mutation
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II. Mutation
11.4 Mutations and Mutants - definitions
11.5 Molecular Basis of Mutation
11.6 Mutation Rates
11.7 Mutagenesis
11.8 Mutagenesis and Carcinogenesis: The Ames
Test
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11.4 Mutations and Mutants - definitions
Mutation
Heritable change in DNA sequence that can lead to a
change in phenotype (observable properties of an organism)
Mutant
A strain of any cell or virus differing from parental strain in
genotype (nucleotide sequence of genome)
Wild-type strain
Typically refers to strain isolated from nature
Animation: The Molecular Basis of MutationsAnimation: The Molecular Basis of Mutations
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11.4 Mutations and Mutants – definitions (cont’d)
Selectable mutations
Those that give the mutant a growth advantage under certain
environmental conditions
Useful in genetic research
Nonselectable mutations
Those that usually have neither an advantage nor a
disadvantage over the parent
Detection of such mutations requires examining a large
number of colonies and looking for differences (screening)
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Selectable and Nonselectable Mutations
Figure 11.4
Selectable mutants: Antibiotic resistance colonies can be detected around a zone of clearance created by the inhibition of a sensitive bacterium
Nonselectable mutants: Aspergilus nidulans produces different interchangeable spontaneously.
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11.4 Mutations and Mutants
Screening is always more tedious than selection
Methods available to facilitate screening
E.g., replica plating
Replica plating is useful for identification of cells with a
nutritional requirement for growth (auxotroph)
Animation: Replica PlatingAnimation: Replica Plating
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Screening for Nutritional Auxotrophs
Figure 11.5
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11.5 Molecular Basis (Types ) of Mutation
Induced mutations
Those made deliberately
Spontaneous mutations
Those that occur without human intervention
Can result from exposure to natural radiation or oxygen radicals
Point mutations
Mutations that change only one base pair
Can lead to single amino acid change in a protein or no
change at all
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Possible Effects of Base-Pair Substitution
Figure 11.6
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11.5 Molecular Basis (consequences) of Mutation
Silent mutation
Does not affect amino acid sequence
Missense mutation
Amino acid changed; polypeptide altered
Nonsense mutation
Codon becomes stop codon; polypeptide is incomplete
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11.5 Molecular Basis of Mutation
Deletions and insertions cause more dramatic
changes in DNA
Frameshift mutations
Deletions or insertions that result in a shift in the reading
frame
Often result in complete loss of gene function
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Shifts in the Reading Frame of mRNA
Figure 11.7
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11.5 Molecular Basis of Mutation
Genetic engineering allows for the introduction of
specific mutations (site-directed mutagenesis)
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11.5 Molecular Basis of Mutation
Point mutations are typically reversible
Reversion
Alteration in DNA that reverses the effects of a prior
mutation
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11.5 Molecular Basis of Mutation
Revertant
Strain in which original phenotype that was changed in
the mutant is restored
Two types
Same-site revertant: mutation restoration activity is at the
same site as original mutation
Second-site revertant: mutation is at a different site in the
DNA
suppressor mutation that compensates for the effect of the
original mutation
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11.6 Mutation Rates
For most microorganisms, errors in DNA replication
occur at a frequency of 10-6to10-7 per kilobase
DNA viruses have error rates 100 – 1,000 X greater
The mutation rate in RNA genomes is 1,000-fold higher
than in DNA genomes
Some RNA polymerases have proofreading capabilities
Comparable RNA repair mechanisms do not exist
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11.7 Mutagenesis
Mutagens: chemical, physical, or biological agents that
increase mutation rates
Several classes of chemical mutagens exist
Nucleotide base analogs: resemble nucleotides
Chemical mutagens can induce chemical modifications
I.e., alkylating agents like nitrosoguanidine
Acridines: intercalating agents; typically cause frameshift
mutations
Animation: MutagensAnimation: Mutagens
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Nucleotide Base Analogs
Figure 11.8
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Chemical and Physical Mutagens and their Modes of Action
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11.7 Mutagenesis
Several forms of radiation are highly mutagenic
Two main categories of mutagenic electromagnetic
radiation
Non-ionizing (i.e., UV radiation)
Purines and pyrimidines strongly absorb UV
Pyrimidine dimers is one effect of UV radiation
Ionizing (i.e., X-rays, cosmic rays, and gamma rays)
Ionize water and produce free radicals
Free radicals damage macromolecules in the cell
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Wavelengths of Radiation
Figure 11.9
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11.7 Mutagenesis
Perfect fidelity in organisms is counterproductive
because it prevents evolution
The mutation rate of an organism is subject to change
Mutants can be isolated that are hyperaccurate or have
increased mutation rates
Deinococcus radiodurans is 20–200 times more
resistant to radiation than E. coli
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11.8 Mutagenesis and Carcinogenesis: The Ames Test
The Ames test makes practical use of bacterial
mutations to detect for potentially hazardous
chemicals
Looks for an increase in the rate of back mutation
(reversion) of auxotrophic strains in the presence of
suspected mutagen
A wide variety of chemicals have been screened for
determining carcinogenicity
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The Ames Test to Assess the Mutagenicity of a Chemical
Figure 11.11
Auxotrophs with single point mutations will not grow in if the required nutrient (eg an amino acid) is not included in the medium. However, in the presence of an added mutagen, some of the cells will revert to wild type an will grow. Eg Histidine-requiring mutants of Salmonella entrica (above)- colonies grow on both plates due to spontaneous mutation but colonies appear on the RHS plate which contains a mutagen)
Disc, no added mutagen Disc, with added mutagen
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DNA REPAIR
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DNA Repair
Three Types of DNA Repair Systems Direct reversal: mutated base is still recognizable and can
be repaired without referring to other strand eg by photoreactivation fromUV damage in which T-T dimers are formed
Repair of single strand damage: damaged DNA is removed and repaired using opposite strand as template eg Excision repair
Repair of double strand damage: a break in the DNA Requires more error-prone repair mechanisms eg SOS repair
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DNA Repair
Pyrimidine dimers form due to exposure to UV radiation (260 nm) – an absorption maxima for DNA .
There are 4 mechanisms by which pyrimidine dimers can be repaired – Refer to htp://trishul.ict.griffith.edu.au/courses/ss12bi/repair.html
Note: Some of the these mechanisms are also used for repairing mutations caused by other mutagenic agents.
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III. Genetic Exchange in Prokaryotes
11.9 Genetic Recombination
11.10 Transformation
11.11 Transduction
11.12 Conjugation: Essential Features 11.13 The Formation of Hfr Strains and Chromosome
Mobilization
11.14 Complementation
11.15 Gene Transfer in Archaea
11.16 Mobile DNA: Transposable Elements
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11.9 Genetic Recombination –definition & mechanism
Genetic Recombination Refers to physical exchange between two DNA
molecules – results in new combination of genes on the chromosome
Ex- fragment aligning, breaking at points, switching & rejoining of alleles of the same gene on two different chromosomes.
Homologous recombination Process that results in genetic exchange between
homologous DNA from two different sources (alleles) (next fig)
Selective medium can be used to detect rare genetic recombinants (fig, after next)
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A Simplified Version of Homologous Recombination
Figure 11.13
Endonuclease nicks one strand of donor DNA, is displaced (eg helicase), & ss binding protein binds. RecBCD has both endonuclease & helicase activities
Strand invasion: RecA (error-prone repair) binds to ss DNA to form a complex & subsequently displaces the complimentary sequence of the other strand to form a heteroduplex (Holliday junction)
Holliday junctions are energised by several proteins & can migrate along the DNA until “resolved” by resolvase – cut & rejon the 2nd & previously unbroken strand
Two types of products of resolvase which differ in conformation can exist in E. coli – patch or splice
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Result of recombination events
Recombination - a recombinant cell is formed Selective medium can be used to detect rare genetic
recombinants
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Recombination and Gene Transfer
But one question still remains...how did the chromosome segment get into the cell for recombination to occur:
The answer is Genetic Transfer!
The players in genetic recombination are:
host cell (host DNA)
donor cell (donor DNA)
DNA is transferred from donor to host (gene transfer)
• Transformation (naked DNA)
• Conjugation (cell to cell contact)
• Transduction (phage mediated)
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11.10 Transformation
Transformation Genetic transfer process by which DNA is incorporated
into a recipient cell and brings about genetic change
Discovered by Fredrick Griffith in 1928 Worked with Streptococcus pneumoniae (see the next
slide to see how he deciphered this process)
This process set the stage for the discovery of DNA
NOTE: Though farmers had known for centuries that crossbreeding of animals
and plants could favor certain desirable traits, Mendel's pea plant experiments
(1856 - 1863) established many of the rules of heredity, now referred to as the
laws of Mendelian inheritance.
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Griffith’s Experiments with Pneumococcus
Figure 11.15 Death due to pneumonia
S=smooth colonies, capsulated, virulent
R = rough colonies, non-capsulated, avirulent
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Streptococcus pneumoniae, phylum Firmicutes causes pneumonia
in mammals. Colonies of the bacteria on petri plates are of two
types:
Smooth due to presence of capsules (polysaccharide) are
virulent and rough (non-capsulated) are avirulent
Cultures from blood samples from dead mice follow Koch's
postulates
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11.10 Transformation
Competent: cells capable of taking up DNA and
being transformed
In naturally transformable bacteria, competence is
regulated
In other strains, specific procedures are necessary to
make cells competent and electricity can be used to
force cells to take up DNA (electroporation)
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11.10 Transformation
During natural transformation, integration of transforming DNA is
a highly regulated, multi-step process
Animation: TransformationAnimation: Transformation
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Mechanisms of Transformation in Gram-Positive BacteriaF
igu
re 1
1.1
6
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11.10 Transformation
Transfection
Transformation of bacteria with DNA extracted from a
bacterial virus
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11.11 Transduction
Transduction Transfer of DNA from one cell to another is mediated by
a bacteriophage. Bacteriophage (phage) are obligate intracellular
parasites that multiply inside bacteria by making use of some or all of the host biosynthetic machinery (i.e., viruses that infect bacteria
Structure of T4 bacteriophage
Contraction of the tail sheath
of T4
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11.11 Transduction
Animation: Generalized TransductionAnimation: Generalized Transduction
There are two types of transduction:
– generalized transduction: A DNA fragment is transferred from one bacterium to another by a lytic bacteriophage that is now carrying donor bacterial DNA due to an error in maturation during the lytic life cycle.
– specialized transduction: A DNA fragment is transferred from one bacterium to another by a temperate bacteriophage that is now carrying donor bacterial DNA due to an error in spontaneous induction during the lysogenic life cycle
Animation: Specialized TransductionAnimation: Specialized Transduction
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11.11 Transduction
Specialized transduction: DNA from a specific
region of the host chromosome is integrated directly
in the virus genome
DNA of temperate virus excises incorrectly and takes
adjacent host genes along with it
Transducing efficiency can be high
Animation: Specialized TransductionAnimation: Specialized Transduction
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Seven steps in Generalised Transduction
1. A lytic bacteriophage adsorbs to a susceptible bacterium.
2. The bacteriophage genome enters the bacterium. The genome directs the bacterium's metabolic machinery to manufacture bacteriophage components and enzymes
3. Occasionally, a bacteriophage head or capsid assembles around a fragment of donor bacterium's nucleoid or around a plasmid instead of a phage genome by mistake.
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4. The bacteriophages are released.
5. The bacteriophage carrying the donor bacterium's DNA adsorbs to a recipient bacterium
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http://www.cat.cc.md.us/courses/bio141/lecguide/unit4/genetics/recombination/transduction/transduction.html
6. The bacteriophage inserts the donor bacterium's DNA it is carrying into the recipient bacterium .
7. The donor bacterium's DNA is exchanged for some of the recipient's DNA.
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Six steps in Specialised Transduction
1. A temperate bacteriophage adsorbs to a susceptible bacterium and injects its genome .
2. The bacteriophage inserts its genome into the bacterium's nucleoid to become a prophage.
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3. Occasionally during spontaneous induction, a small piece of the donor bacterium's DNA is picked up as part of the phage's genome in place of some of the phage DNA which remains in the bacterium's nucleoid.
4. As the bacteriophage replicates, the segment of bacterial DNA replicates as part of the phage's genome. Every phage now carries that segment of bacterial DNA.
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5. The bacteriophage adsorbs to a recipient bacterium and injects its genome.
6. The bacteriophage genome carrying the donor bacterial DNA inserts into the recipient bacterium's nucleoid.
http://www.cat.cc.md.us/courses/bio141/lecguide/unit4/genetics/recombination/transduction/spectran.html
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Summary – specialized transduction
DNA from a specific region of the host chromosome is integrated directly in the virus genome
A of temperate virus excises incorrectly and takes adjacent host genes along with it
Transducing efficiency can be high
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11.12 Conjugation: Essential Features
Bacterial conjugation (mating): mechanism of
genetic transfer that involves cell-to-cell contact
Plasmid encoded mechanism
Donor cell: contains conjugative plasmid
Recipient cell: does not contain plasmid
Animation: ConjugationAnimation: Conjugation
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11.12 Conjugation: Essential Features
F (fertility) plasmid
Circular DNA molecule; ~ 100 kbp
Contains genes that regulate DNA replication
Contains several transposable elements that allow the
plasmid to integrate into the host chromosome
Contains tra genes that encode transfer functions
Animation: Conjugation FAnimation: Conjugation F
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Genetic Map of the F (Fertility) Plasmid of E. coli
Figure 11.19
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11.12 Conjugation: Essential Features
Sex pilus is essential for conjugation
Only produced by donor cell
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Formation of a Mating Pair
Figure 11.20
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11.12 Conjugation: Essential Features
DNA synthesis is necessary for DNA transfer by
conjugation
DNA synthesized by rolling circle replication;
mechanism also used by some viruses
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Transfer of Plasmid DNA by Conjugation
Figure 11.21a
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Transfer of Plasmid DNA by Conjugation
Figure 11.21b
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11.13 The Formation of Hfr Strains and Chromasome Mobilization
F plasmid is an episome; can integrate into host
chromosome
Cells possessing a non-integrated F plasmid are called
F+
Cells possessing an integrated F plasmid are called Hfr
(high frequency of recombination)
High rates of genetic recombination between genes on
the donor chromosome and those of the recipient
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Presence of the F plasmid results in alterations in
cell properties
Ability to synthesize F pilus
Mobilization of DNA for transfer to another cell
Alteration of surface receptors so that cell can no longer
act as a recipient in conjugation
11.13 The Formation of Hfr Strains and Chromasome Mobilization
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Insertion sequences (mobile elements) are present
in both the F plasmid and E. coli chromosome
Facilitate homologous recombination
11.13 The Formation of Hfr Strains and Chromasome Mobilization
Animation: Conjugation HfrAnimation: Conjugation Hfr
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The Formation of an Hfr Strain
Figure 11.22
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Transfer of Chromosomal Genes by an Hfr Strain
Figure 11.23
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11.13 The Formation of Hfr Strains and Chromosome Moblilization
Recipient cell does not become Hfr because only a
portion of the integrated F plasmid is transferred by the
donor
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Transfer of Chromosomal DNA by Conjugation
Figure 11.24
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Hfr strains that differ in the integration position of the F
plasmid in the chromosome transfer genes in different
orders
11.13 The Formation of Hfr Strains and Chromosome Moblilization
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Formation of Different Hfr Strains
Figure 11.25
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Identification of recombinant strains requires selective
conditions in which the desired recombinants can grow
but where neither of the parental strains can grow
11.13 The Formation of Hfr Strains and Chromosome Moblilization
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Example Experiment for the Detection of Conjugation
Figure 11.26
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Genetic crosses with Hfr strains can be used to map
the order of genes on the chromosome
11.13 The Formation of Hfr Strains and Chromosome Moblilization
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Time of Gene Entry in a Mating Culture
Figure 11.27
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11.13 The Formation of Hfr Strains and Chromosome Mobilization
F′ plasmids
Previously integrated F plasmids that have excised and
captured some chromosomal genes
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11.14 Complementation
Merodiploid (or partial diploid)
Bacterial strain that carries two copies of any particular
chromosomal segment
Complementation
Process by which a functional copy of a gene
compensates for a defective copy
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11.14 Complementation
Complementation tests are used to determine if two
mutations are in the same or different genes
Necessary when mutations in different genes in the same
pathway yield the same phenotype
Two copies of region of DNA under investigation must be
present and carried on two different molecules of DNA (trans
configuration)
Placing two regions on a single DNA molecule (cis
configuration) serves as a positive control for these tests
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Complementation Analysis
Figure 11.28
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11.14 Complementation
Cistron: gene defined by cis-trans test
Equivalent to defining a structural gene as a segment
of DNA that encodes a single polypeptide chain
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11.15 Gene Transfer in Archaea
Development of gene transfer systems for genetic
manipulation lag far behind Bacteria
Archaea need to be grown in extreme conditions
Most antibiotics do not affect Archaea
No single species is a model organism for Archaea
Examples of transformation, viral transduction, and
conjugation exist
Transformation works reasonably well in Archaea
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An Archaeal Chromosome Viewed by Electron Microscope
Figure 11.29
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11.16 Mobile DNA: Transposable Elements
Discrete segments of DNA that move as a unit from one
location to another within other DNA molecules (i.e.,
transposable elements)
Transposable elements can be found in all three domains
of life
Move by a process called transposition
Frequency of transposition is 1 in 1,000 to 1 in 10,000,000
per generation
First observed by Barbara McClintock
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11.16 Mobile DNA: Transposable Elements
Two main types of transposable elements in
Bacteria are transposons and insertion sequences
Both carry genes encoding transposase
Both have inverted repeats at their ends
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Maps of Transposable Elements IS2 and Tn5
Figure 11.30
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11.16 Mobile DNA: Transposable Elements
Insertion sequences are the simplest transposable
element
~1,000 nucleotides long
Inverted repeats are 10–50 base pairs
Only gene is for the transposase
Found in plasmids and chromosomes of Bacteria and
Archaea and some bacteriophages
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11.16 Mobile DNA: Transposable Elements
Transposons are larger than insertion sequences
Transposase moves any DNA between inverted repeats
May include antibiotic resistance
Examples are the tn5 and tn10
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11.16 Mobile DNA: Transposable Elements
Mechanisms of Transposition: Two Types
Conservative: transposon is excised from one location
and reinserted at a second location (i.e., Tn5)
Number of transposons stays constant
Replicative: a new copy of transposon is produced and
inserted at a second location
Number of transposons present doubles
Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Transposition
Figure 11.31
Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Two Mechanisms of Transposition
Figure 11.32
Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings
11.16 Mobile DNA: Transposable Elements
Using transposons is a convenient way to make mutants
Transposons with antibiotic resistance are often used
Transposon is introduced to the target cells on a plasmid that
can’t be replicated in the cell
Cells capable of growing on selective medium likely acquired
transposon
Most insertions will be in genes that encode proteins
You can then screen for loss of function and determine
insertion site
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Transposon Mutagenesis
Figure 11.33