Bacterial Genetics

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Bacterial Genetics
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Review

• Genome genetic blueprint
• Gene
• Most organisms-DNA
• Viruses
• DNA or RNA

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Nucleic acids

• Nucleotides
• Sugar
• Phosphate
• Nitrogenous base
• DNA
• Guanine-G cytosine-C adenine-A thymine-T
• RNA
• Uracil-U replaces thymine

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Review

• DNA ds helix
• Strands held together by H bonds
• Complimentary base pairing
• A pairs with T in RNA, A pairs with U
• C with G
• Linear sequence of bases contains info
• 3 nucleotides code for 1 amino acid

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Duplication of DNA

• Replication of chromosome(s)
• Precise duplication of DNA
• DNA polymerase
• Denaturation separate strands

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Terms

• Genotype set of genes carry instructions
• Phenotype expression of those genes
• Genome of prokaryotes includes chromosomal DNA
  and plasmids

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Flow of Information

• DNA to RNA to polypeptides
• Transcription-synthesis of complimentary strand
  of RNA from DNA-mRNA
• Separates strands of DNA
• Copies only DNA needed for protein synthesis-mRNA

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Flow of Information

• Translation protein synthesis
• Decodes sequence of nucleotides into amino acids
  (20)
• Genetic Code codons, group of 3
  nucleotides-AAU,CGC
• AUG-start codon

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Degenerate Code

• Each codon specifies a particular amino acid
• Several codons code same aa

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Flow of Information

• Translation
• Ribosomes-site of protein synthesis
• tRNA recognizes the specific codon
• tRNA forms complimentary base pairing with the
  codon
• Has anticodon carries the required aa

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Question

• AUG is first codon on mRNA.
• What is the anticodon on the tRNA?

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Regulation of Gene Expression

• Prokaryotes Where is mRNA transcribed in cell?
• Translation can begin before completion of mRNA
  molecule

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Regulation of Gene Expression

• Eukaryotes Where?
• mRNA contains exons and introns
• Exons are expressed, encode for aa
• Introns do not
• Processing by ribozymes to remove introns
  splice exons

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Regulation of Gene Expression

• Conserve energy
• Turn on off transcription of genes
• Constitutive genes not regulated
• Operon

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Operons

• Regulation of metabolic genes
• Uses repressors (regulatory genes)
• Block RNA polymerase from attaching

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Induction

• Turn on the transcription of gene
• Inducer- induces transcription
• Inducible enzymes
• Synthesized only when substrate is present
• Glycolysis genes constitutive

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Lac Operon

• Inducible operon enzymes to metabolize lactose
• Default is off
• Regulatory sites
• Promoter-
• Operator-

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Lac Operon

• i genes code for repressor-regulatory protein
• Always turned on
• Binds to operator
• Structural genes
• Lac operon-3 genes

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Lactose in Medium

• Binds repressor changing shape
• Repressor cant bind
• RNA polymerase can bind
• Enzymes for lactose metabolism produced
• To turn on operon

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Repressible Operon

• Tryptophan operon
• Usually occurs in anabolism
• Repressor is inactive so tyrptophan is
  synthesized
• Default in on

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Tryptophan Operon

• Amino acid in media
• Binds to repressor activating it
• Genes to synthesize amino acid produced

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Mutation

• Change (heritable) in base sequence of DNA
• Called mutant
• Genotype differs from parent
• Phenotype may be altered

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Types of Mutations

• Spontaneous mutations
• DNA replication errors
• Occur at low frequencies
• Induced mutations-mutagens
• Alter structure of bases
• Errors in base paring

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Point Mutations

• Change in 1 base-pair
• Single base is replaced with another
• Change in genotype
• May be change in phenotype

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Types of Point Mutations

• Silent mutation- no phenotypic change
• Degenerate code

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Missense Mutations

• Change in amino acid
• Can result in significant changes in polypeptide

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Examples of Missense

• Sickle cell anemia- hemoglobin
• Change from glutamic acid (hydrophilic) to valine
  (hydrophobic)
• Change in shape of protein

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Nonsense Mutation

• Base-pair substitution
• Create stop codon in middle of mRNA
• Premature termination of translation

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Frame Shift Mutation

• Bases deleted or inserted
• Shifts translational reading frame
• Large insertions are transposons

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Frame Shift

• THE FAT CAT ATE THE BAD RAT
• Remove a C
• THE FAT ATA TET HEB ADR AT

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Induced Mutations

• Mutagens
• Increase mutation rate
• Chemical mutagens
• Nitrous acid
• Occurs at random sites

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Radiation

• UV light -nonionizing
• Covalent bonds between certain bases
• Adjacent thymines(Ts) can cross link
• Pyrimidine dimers
• Some enzymes that repair UV damage

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Nucleoside Analog

• Structurally similar to normal bases
• Have altered base pairing
• 2 aminopurine replaces A may pair with C
• 5-bromouracil replaces T but may pair with G

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Genetic Recombination

• Physical exchange of genes between 2 homologous
  DNA molecules
• Contributes to populations genetic diversity

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Horizontal Transfer

• Microbes of same generation
• Involves a donor cell - gives DNA to recipient
  cell
• Part of DNA incorporated into recipients DNA

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Transformation

• Free (naked) DNA in solution
• Cells after death, release DNA
• Cells may take up DNA
• Only in certain stage of cell cycle

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Competence

• Cells able to take up DNA be transformed
• Release competence factor that helps in uptake

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Transformation

• Enzymes cut DNA into small pieces
• Recombination between donor recipient
• Few competent bacteria

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First Evidence of Transformation

• Griffith in 1920s
• Streptococcus pneumoniae in mice

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Conjugation

• Mediated by one kind of plasmid
• F plasmid or F factor
• Genes to control conjugation
• Donor cells must have F plasmid

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Differs from Transformation

• Cells must be of opposite mating types
• Donor is F plus
• Recipient is F minus
• Requires direct cell contact
• Transfers larger quantities of DNA

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Conjugation

• Gram negative cells
• Gram positive cells produce sticky surface
  molecules
• Keeps cells together

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Conjugation

• Plasmid is replicated
• A copy of plasmid transferred to recipient
• F minus cell becomes F plus
• Receptors on new F plus change

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F Plasmid

• Plasmid integrates into the chromosome converts
  cell to Hfr cell( high frequency of
  recombination)
• F factor DNA can separate and become plasmid

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Conjugation

• Hfr and F- cell
• Replication of Hfr begins in middle of integrated
  F factor
• Small piece leads the chromosome into F- cell
• Donor DNA can recombine (DNA not integrated is
  degraded)

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Transduction

• Bacterial DNA is transferred via a virus
• -Bacteriophage
• Virulent phages lytic cycle
• Generalized transduction
• Any gene on donor chromosome transferred

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Generalized Transduction

• All genes are equally likely to be packaged
  inside phage
• Virus cannot replicate in new bacteria
• Defective virus
• Specialized transduction-only certain bacterial
  genes transferred

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Plasmids

• Extra chromosomal material
• F factor or plasmid is a conjugative plasmid
• Carries genes for sex pili and for transfer

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Dissimilation Plasmids

• Enzymes break down unusual sugars and
  hydrocarbons
• Pseudomonas use toluene and petroleum as carbon
  and energy sources
• Used to clean up oil spills
• Allows bug to grow in adverse environments

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Virulence Plasmids

• E. coli carries plasmids that code for
• toxins diarrhea
• Bacteriocins- toxic proteins kills other bacteria
• E. coli produces colicins

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Resistance Factors

• R factors- resistance to antibiotics, heavy
  metals or cellular toxins
• Wide spread use of antibiotics- led to selection
  of bacteria with R factors with resistant genes

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Transposition

• Segment of DNA moves from one place in chromosome
  to another
• Rare event
• Transposable elements
• Insert within a gene inactivate it

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Transposons

• Contain information for own transposition
• Insertion sequences (IS) contain only gene for
  transposase
• Complex transposons carry genes for
  enterotoxins or antibiotic resistance