Microbial GeneticsWHY?termsGenotype refers to genetic makeupPhenotype refers to expression of that genetic makeupHeritable traits must be encoded in DNAMutations occur randomly

Mutants resistant to an antibiotic

Kinds of mutantsAuxotroph: loss of biosynthetic enzymeCold- or temperature-sensitive: enzyme function is restricted to narrower temperature rangeDrug resistance or sensitivity: permeability, degradation, or site of actionMorphology: change in capsule or other feature leads to change in colonyCatabolic: loss of a degradative enzyme leads to, for example, inability to ferment a carbohydrateVirus-resistant: loss of surface feature that is a virus receptor

Results of point mutations

Frame-shift mutations

ReversionsSame-site reversion: may be true revertant (same sequence) or just restore activitySecond-site reversionFrame-shiftProduction of another enzyme that fulfills the function

MutagensAnalogs for bases5-bromo-uracil for thymine (5BU can pair with G as well as with A)2-aminopurine for adenine (2AP can pair with C as well as with T)Intercalating agents (ethidium bromide)UV (260 nm) causes dimerization of adjacent thymines (photoreactivation)Ionizing radiation causes chemical changes to the DNAGeneration of free radicals that can react with DNASS or DS breaks in the DNA moleculeTransposable genetic elements

Recombination

Recombination

Genetic exchange: transformation

Genetic exchange: plasmid transferSmall, usually circular, independently replicating DNA moleculesGenerally, G- plasmids replicate as does chromosomal DNA, G+ plasmids by rolling circle replicationGenes of replication control, timing initiation on plasmid (ori)Some plasmids integrate (F+, Hfr)Most are double-strandedAbout 1- 100kbCode for:R-factors (R-plasmids) : antibiotic resistance, heavy metal resistanceVirulence plasmids : adhesins, hemolytic factors, toxin, Ti, bacteriocinsDegradation, tol, nah,Plasmid copy #Compatibility (inc)

Avery Experiment

TransformationCompetence: ability to be transformedSteps in transformationDNA binding and uptake (SS or DS, depending on species)Integration (recA)Competence may be induced by electroporation, Ca

Mechanism of transformation

TransductionGeneralizedcan be carried out by either lytic or temperate phageSpecializedrequires specific integration

Generalized transduction

Specialized transduction

Specialized transduction

Phage ConversionLysogeny may cause other changes in the host cellOften the host acquires immunity to additional infection by that phage typeThere may be other changes that may be beneficial to the hostLysogenized Salmonella anatum acquires cell-surface changesLysogenized Corynebacterium diphtherium acquires toxin

Genetic exchange: conjugation

Conjugation: earlyCell-surface structure

Conjugation: middle

Conjugation: late

TransposonsRare eventsMobile Genetic elements-jumping genesCarry a transposase, and flanked by inverted repeats20bp to >100bpFirst discovered in maize/cornPhase variation (invertible elements, need invertase)

Transposons

Genetic Engineering-basicsBasic steps in cloningRestriction-modification enzymesshot gun or PCRLigaseRecombination Plasmids (or phage)= vectorsExpression vectorsSelection of clonesLooking for a clone with a specific geneProbes (DNA, RNA or antibody)Wave of the future: DNA chips or microarrays, BAC libraries, automated sequencing etc

GenomicsBioinformaticsHarvesting genes for biotech (Diversa)Recent Science articleComparing gene families

Escherichia coli map

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