Microbial Genetics Flashcards
Mutant
strain of any cell/virus differing from the parental strain in genotype
Selectable Mutations
give mutants a growth advantage under certain conditions
Nonselectable Mutations
have neither an advantage nor a disadvantage over the parent
Induced Mutations
mutations made environmentally (from natural radiation or oxygen radicals) or deliberately
Spontaneous Mutations
mutations that occur w/o external intervention; inherent mistakes made by polymerases
Point Mutations
mutations that change only 1 bp; can lead to silent; missense; or nonsense mutations
Frameshift Mutations
deletions or insertions that result in shift in reading frame; often result in complete loss of gene fxn
Transition Mutation
purine base is mutated to a purine OR pyrimidine base is mutated to a purimidine
Transverison Mutation
purine base is mutated to a pyrimidine OR pyrimidine base is mutated to a purine
Are All Mutations Equally Harmful?
No
How harmful are mutations in tRNAs?
They may be highly detrimental or not.
How harmful are mutations in rRNAs and other functional RNAs?
They are less detrimental than mutations in tRNA
How harmful are point mutations in protein-coding genes?
Those in catalytic sites are very harmful.
Those in structural sites are not as harmful.
How harmful are frameshift mutations in protein-coding genes?
Those early in a sequence are very harmful.
Those late in a sequence are less harmful.
What is the mutation rate for errors in DNA replication for most microorganisms?
The frequency is 10-6 - 10-7
How do the mutation rates of DNA viruses compare to most other microorganisms?
The mutation rate is 100-1000x greater than the microorganisms’ (10-6 - 10-7).
How does the mutation rates of RNA genomes compare to the mutation rates of DNA genomes?
RNA genomes have mutation rates that are 1000-fold higher because similar repair mechanisms to the ones in DNA do not exist.
Mutagens
chemical, physical, or biological agents that increase mutation rates
Nucleotide Base Analogs - Mutagens
mutagens that resemble nucleotides and that cause mispairing
Chemical Mutagens
chemical mutagens can…
- modify bases = mispair
- cause bubble in DNA during replication = lethal ->
- to fix…
- pol falls off and proteins come in that insert/delete nucleotides = frameshift
- to fix…
Non-Ionizing Mutagenic Radiation
- UV Radiation
- purines and pyrimidines strongly absorb UV
- can cause pyrimidine dimer (ex. T=T)
Ionizing Mutagenic Radiation
- X-rays, cosmic rays, gamma rays
- ionize water and produce free radicals which damage macromolecules in cell
What are 3 classes of ways to repair mutations?
- Single Strand Break Repair
- Double Strand Break Repair
- Direct Reversal Break Repair
What is single strand break repair and its 3 types?
A repair where the damaged DNA is removed and repaired using the opposite strand as template
- Mismatch Repair
- Nucleotide Excision Repair
- Base Excision Repair
What is double strand break repair and its types?
A repair of a break in DNA by filling in bases randomly = error-prone
- Recombinational Repair
What is direct reversal repair and its types?
A repair where the mutated base is still recognizable and can be repaired w/o referring to the other strand
- Error Prone Repair
- Error Free Repair
Steps of the Error Free Repair Mechanism:
- Lesions detected by RNA pol - recruits Mfd, recruits UvrA
- RNA pol released, UvrB is recruited
- Lesion is verified
- UvrB makes pre-incision complex
- UvrB recruits UvrC, which makes dual incision
- Excision of mutation by UvrD
- DNA pol and ligase rebuild the excised sequence
Which repair system is used when the damage is large scale?
SOS Response
What is the SOS Response?
The SOS Response is a regulon which regulates over 40 genes that are involved in many DNA repair systems.
Steps of the SOS Response Mechanism:
- DNA damage activates RecA
- RecA activates LexA’s (repressor of SOS) protease activity which then cleaves itself
- w/o the repression from LexA the regulated genes become highly active with their repair systems
What is the error prone repair system?
The Translesion Synthesis which allows the DNA to be repaired/synthesized w/ no template.
Steps of the Translesion Synthesis Mechanism:
- pol 3 reaches error translesion and falls off
- replaced by pol 4 and pol 5 which insert any base into the translesion
- other enzymes can come and fix base mispairing
What is an example of differences in mutation rates for mutants between organisms?
Deinococcus radiodurans is 20-200x more resistant to radiation than E.coli
To which phylum does the Water Bear belong?
The Tartigrades phylum
Technically; Water Bears are what?
micro-animals
Water Bears are resistant to what?
They are resistant to …
- temps just above 0 K (extremophiles)
- temps well above boiling
- high pressures
- radiation (1000x more resistant than humans)
- nutrient lim. for >100 yrs
- dessication
What are the 3 methods for genetic exchange in bacteria?
- Transformation
- Conjugation
- Transduction
What is consistent with all of the methods for bacterial genetic exchange?
They all rely on recombination to incorporate DNA.
What is Genetic Recombination?
The physical exchange of DNA between genetic elements.
What is Homologous Genetic Recombination?
Recombination that occurs between regions of similar DNA
Steps of Homologous Genetic Recombination:
- endonuclease nicks donor DNA
- binding of SSB protein to the nicked site
- strand invasion of recipient DNA bound with RecA protein
- cross-strand exchange occurs
* creates both patches and splices
What is Transformation?
The genetic transfer process by which DNA, often from the environment, is incorporated into recipient cell and brings about genetic change.
Who discovered transformation?
Fredrick Griffith in late 1920s
What does Competency mean?
The capability of cells to take up naked DNA through transformation.
- need DNA binding protein, autolysins (for peptidoglycan), nucleases
Are all cells naturally competent?
No
- for some cell to be competent they must be forced through procedures using electricity and/or chemicals.
What is an example of a naturally competent bacteria?
B.subtilis
Steps of Transformation:
- DNA-binding protien binds external DNA
- Nucleases cut DNA
- Competence-specific, ssDNA-binding protein bring in ssDNA
- recA mediates the external DNA’s homologous recombination into the recipient cell’s DNA
What is Conjugation?
Mechanisms of genetic transfer that involved cell-to-cell contact; usually only plasmids; from Donor Cell (w/ certain plasmid) to Recipient Cell (w/o certain plasmid)
Steps of Conjugation:
- sex pilus is formed on donor cell (tra genes)
- DNA is synthesized by rolling circle replication to make copy
- DNA copy is transferred through sex pilus
- Sex pilus is degraded
Steps of Rolling Circle Replication:
- DNA strand is nicked by nickase at the origin (OriT)
- Strands are peeled apart by TraI
- Outside strand is transferred out
- Inside strand is template for new strand
What is Transduction?
Transfer of DNA from 1 cell to another by bacteriophage
What are the 2 modes of Transduction?
Generalized Transduction and Specialized Transduciton
What is Generalized Transduction?
Transduction where the DNA,derived from any portion of host genome, is inside mature virion
- mainly done by Lytic viruses
- low efficiency
Steps of Generalized Transduction:
- Host Cell is infected by lytic virus (phage DNA -> host DNA)
- Host cell makes many normal phage, but also rare Transducing Particle
* has no phage DNA, only host DNA - Transducing Particle infects recipient cell (host DNA -> recipient DNA)
* integration through homologous recombination
What is Specialized Transduction?
Transduction of DNA, from specific region of host chromosome, is integrated directly into virus genome
- mainly done by Lysogenic viruses
- high efficiency
Steps of Specialized Transduction:
- Lysogenic virus infects host cell incorrectly, so that the host DNA is incorporated into phage DNA
- Incorrect DNA is replicated inside cell
- Defective phage are made and are released to transduce other cells
Do Archae have mechanisms of gene transfer similar to bacteria?
Yes, there are examples of transformation, transduction, and conjugation.
How do Eukaryotic microbes undergo gene transfer?
They undergo conjugation using ciliates.
- haploid nuclei exchanged in both directions
What are Transposable Elements?
Discrete segments of DNA that move as a unit from 1 location to another w/in other DNA molecules
How do transposable elements move?
Through a process called Transposition.
What are the 2 main types of transposable elements in bacteria?
Transposons
Insertion Sequences
What are Insertion Sequences?
- simplest transposable elements
- ~1000 nucleotides long
- have inverted repeats (10-50 bps)
- only have transposase gene
What are Transposons?
- larger than insertion sequences
- ends are IRs
- somewhere in middle are transposase genes
