Genetics b
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Genetics b

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Genetics b Genetics b Presentation Transcript

  • DNA as the Genetic Material The early work of Fred Griffith in 1928 on the transfer of virulence in the pathogen S. pneumoniae set the stage for DNA research.
  • Transformation
    • Change of nonvirulent bacteria into virulent pathogens.
  • The Genetic Code
    • DNA base sequence corresponds to the amino acid sequence of the polypeptide specified by the gene.
    • Organization of the code – codons, wobble
    • Gene structure – linear sequence of nucleotides
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  • Mutations
    • Spontaneous mutations arise occasinally in all cells and develop in the absence of any added agent.
    • Induced mutations are the result of exposure of the organism to some physical or chemical agent called a mutagen.
  • Biochemical Mutations
    • Prototrophs – can grow on minimal media
    • Resistant mutant – is a particular type of biochemical mutant that acquires resistance to some pathogen, chemical, or antibiotic.
    • Easy to select for and very useful in microbial genetics.
  • Biochemical Mutations
    • Cause a change in the biochemistry of the cell.
    • Inactivate a biosynthetic pathway
    • Auxotrophs – cannot grow on minimal medium
  • Mutations
    • Mutare – to change
    • Conditional mutations – expressed only under certain environmental conditions.
  • Point Mutations
    • Silent
    • Missense
    • Nonsense
    • Frameshift
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  • Suppressor Mutation
    • Overcomes the effect of the first mutation.
    • The second mutation is within the same gene, the change may be called a second site reversion or intergenic suppression.
  • The Expression of Mutations
    • The expression of a mutation will only be noticed if it produces a detectable, altered phenotype.
    • Wild type  forward mutation
    • Reversion mutation  back mutation
  • Spontaneous Mutations
    • Arise from several potential sources.
    • Errors in DNA replication, damage to DNA from factors such as gamma radiation and heat, or transposons.
    • Transition and transversion mutations.
    • Frameshifts – deletion of DNA segments resulting in an altered codon reading frame
  • Induced Mutations
    • Virtually any agent that directly damages DNA, alters its chemistry, or interferes with repair mechanisms will induce mutations.
    • Modes of mutagen action – incorporation of base analogs, specific mispairing, intercalation, and bypass of replication.
  • Mutant Selection
    • Replica plating – direct and indirect method.
    • Ames Test – developed in the 1970s by Bruce Ames, used to detect carcinogens
  • Detection and Isolation of Mutants
    • In order to study microbial mutants, one must be able to detect them readily, even when there are few, and then efficiently isolate them from the parent organism and other mutants.
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  • Microbial Genetics: Recombination and Plasmids
    • Recombination is the process in which a new recombinant chromosome, one with a genotype different from either parent, is formed by combining genetic material from two organisms.
  • Bacterial Plasmids
    • Conjugation, the transfer of DNA between bacteria involving direct contact, depends on the presence of an extra piece of circular DNA – plasmid DNA.
  • Plasmids
    • Are small circular DNA molecules that can exist independently of host chromosomes and are present in many bacteria (some yeast and fungi).
    • Own replication origins and are autonomously replicating and stably inherited.
  • Replicons
    • Is a DNA molecule or sequence that has a replication origin and is capable of being replicated.
    • Plasmids and bacterial chromosomes are separate replicons.
    • Plasmids – less than 30 genes, single or multiple copies
  • Curing
    • Plasmids can be eliminated from host cells in a process known as curing.
    • May occur spontaneously or be induced by treatments that inhibit plasmid replication.
    • Acridine mutations, UV and ionizing radiation, thymine starvation, and growth above optimal temperature.
  • Classified: mode of existence and spread
    • Episome – is a plasmid that can exist either with or without being integrated into the host’s chromosome.
    • Conjugative – plasmids have genes for pili and can transfer copies of themselves to other bacteria during conjugation
  • Fertility Factors
    • A plasmid called the fertility factor or F factor plays a major role in conjugation in E. coli .
    • About 94.5 kilobases long and bears genes responsible for cell attachment and plasmid transfer between specific bacterial strains during conjugation.
  • F factor
    • Also contains insertion sequences that assist plasmid integration into host cell chromosome.
    • The F factor is an episome that can exist outside the bacterial chromosome or be integrated into it.
  • Resistance Factors
    • R factors – often confer antibiotic resistance on the bacteria that contain them
    • Code for enzymes capable of destroying or modifying antibiotics.
    • Usually not integrated into host chromosome
    • Resistance genes are within a transposon
  • Col Plasmids
    • Bacteria also harbor plasmids with genes that may give them a competitive advantage in the microbial world.
    • Bacteriocins – bacterial proteins that destroy other bacteria
    • Virulence - make the bacteria more pathogenic
    • Metabolic – degrade substances
  • Transposable Elements
    • Move around the genome – transposition
    • Insertion sequences – IS elements (750 to 1,600 base pairs) contain only the genes for those enzymes required for its transposition
    • Composite transposons
  • Bacterial Recombination
    • Transformation
    • Transduction
    • Conjugation
  • DNA Transformation
    • Uptake of naked DNA
    • Transformation frequency of very competent cells is around 10 -3 for most genera when an excess of DNA is used
  • Griffith’s Experiment
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  • Transformation
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  • Bacterial Conjugation
    • F + x F -
    • HFr
    • F’
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  • Hfr Conjugation
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  • Transduction
    • Bacterial viruses participate in this mode of bacterial gene transfer.
    • Generatlized transduction
    • Specialized transduction
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  • Generalized Transduction
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  • Specialized Transduction
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  • Altered Porin
  • Altered Carrier Protein
  • Pumps
  • Restriction Endonucleases
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