Download Lecture 9 - Bacterial Genetics Chpt. 8

2/10/2011
Chapter 8: Bacterial Genetics
What are mutations?
• Change in the base sequence of the
DNA
• Do they always change the genetic
code?
Vertical Gene transfer
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Horizontal gene transfer
Spontaneous Mutation
• Spontaneous mutations occur in natural
environment
– Occur infrequently and randomly
• Rate of mutation
– Probability that a mutation will be observed
in a given gene each time the cell divides
– Rate is generally between 1 in 10,000 and
1 in a trillion
• Low rate is due to cellular repair mechanisms
Spontaneous Mutation
• Mutations are stable heritable changes
in the base sequence of DNA
• Can occur from
– Base substitutions
– Removal or addition of nucleotides
– Transposable elements
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Types of Mutations
• Base-pair mutation
– Missense mutation
– Nonsense mutation
– Silent mutation
• Frameshift mutation
• Transposons
Base-pair mutation:
missense
Results of base-pair mutations
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Types of Mutations
• Frameshift
– Changes the reading frame
Frameshift mutation
Types of Mutations
• Transposable elements
– Special segments of DNA that move
spontaneously from gene to gene
• Elements called transposons
– Transposons disrupt proper function
of gene
• Gene or gene product generally nonfunctional
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Barbara McClintock: “jumping
genes” biological mutagen
What can cause mutations?
• Chemicals (nitrous acid)
• Physical mutagens (uv light)
• Biological mutagens (transposons)
Nitrous Acid as a Chemical
Mutagen
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Nucleoside Analogs are
Chemical Mutagens
Intercalating Agents as
Chemical Mutagens
• Molecules that insert themselves
between adjacent bases
–Create space between bases
»Extra base is often added to fill
space
• Ethidium bromide is common
intercalating agent
–Potential carcinogen
Intercalating Agents
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UV Light as a Physical Mutagen
• Radiation
– Two types
• Ultraviolet light
– Causes covalent bonding between adjacent thymine bases
» Forms thymine dimers which distorts DNA
• X-rays
– Cause breaks and alterations in DNA
» Breaks that occur on both strands are often lethal
Repair mechanisms
• Repair of base substitution
– Cells develop two methods of repair
• Proofreading
– DNA polymerase has proofreading function
– Able to excise incorrect base and replace with correct
one
• Mismatch repair
– Endonuclease enzyme removes short stretch of
nucleotide
– DNA polymerase fills gap
– DNA ligase joins ends
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Repair: Mismatch
Repair of UV Damage
• Repair of thymine dimers
– Two mechanisms
• Light repair
– a.k.a photoreactivation
– Enzyme uses visible light to break covalent bonds
between bases
• Dark repair
– a.k.a excision repair
– Endonuclease excises damaged section
– New section replicated and joined to original strand
Light Repair
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Dark Repair
Repair of Damaged DNA
• Repair of modified bases
– Enzyme cuts DNA backbone and removes
base
– DNA polymerase incorporates new base
• SOS repair
– Last ditch effort to bypass damage
– Damage induces SOS system
• Produces new DNA polymerase
– Highly error prone
» Mutations can arise from synthesis with new
enzyme
Repair of Damaged DNA
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Why use bacteria to study
mutations?
• Only have one chromosome…one copy
of each gene
• Easy to grow
Direct Selection
• Testing for traits that are easily
identified
– Resistance to antibiotics
– Motility
Direct selection
Involves inoculating population
of bacteria on medium on which
only mutants will grow
Used to select antimicrobialresistant organisms
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Indirect Selection
• A way to look at traits that are not easily
identified, e.g. changes in metabolic
p
pathways
y
• Replica plating
– A way to identify AUXOTROPHS from
PROTOTROPHS
Indirect selection
Required to isolate organisms that
require growth factor that parent
strain does not
Replica plating
Testing chemicals for mutagenicity…Ames test
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