Lambda phage

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Lambda phage

• ??? ?????????????? ???????????? 45040052

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Lambda phage

• Order Caudovirales
• Family Siphoviridae
• Genus ?-like viruses
• Species Enterobacteria phage ?

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Morphology

• Head icosahedral symmetry
• Tail Helical Symmetry
• one tail fiber
• Capsid not enveloped
• Linear dsDNA (phage)
• Infected E.coli cell

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Genome

• genome contains about 50,000 nucleotide pairs
• encodes 50-60 different proteins
• Genome is 54 of virion by weight

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• The ends of the genome have sticky ends 12 bp
  long gt cos sites (cohesive ends)
• The ends are joined by bacterial enzyme, Ligase
• Closed circle dsDNA (E.coli)

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linear genome becomes circularized
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Infected in E.coli cell
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Life cycle of lambda phage
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Lytic and Lysogenic phage

• Lytic pathway
• - ????????????????????????????????
• - ?????????? (cell lysis)
• - Cro promotes the lytic phase
• Lysogenic pathway
• - ?????????????????? ???? prophage
• ???? induction event ???????
• lytic pathway
• - CI promotes the lysogenic cycle
• "right" operater of lambda

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Lytic cycle

• 1. Adsorption specific with
  surface of the cell (outer memb.)
• 2. Injection
• 3. Circulation cos site
  ,nuclease host
• 4. Replication
  bidirectional
• 5. Protein coding
  endonuclease plus st.
• 6. Syn new minus strand and
  new plus st.
• 7. Lytic rolling circle
  long DNA for multiple phage genome
• 8. Code structeral protein ??? DNA ??? host ???
  phage
• 9. Packaging
• 10. Phage code endolysin destroy peptidoglycan
• Cell lysis

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Lytic cycle

• Nick
• Rolling circle
• Polymeric genome
• Endonuclease
• Cohesive end

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Lysogengic cycle

• ???????????
• C1 protein ???? repressor protein ??? Cro
  protein ??? C1 protein ?????????????????????
  promotor ?????????? Cro promotor ????? Inactivate
  Cro promotor ??????????????? Lytic cycle

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Lysogengic cycle

• Phage DNA is integrated into host chromosome at
  attP site in phage genome and attB site in host
  genome

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Integration

• att an E.coli seqence for the "attachment" or
  integration of lambda's circular chromosome.
• oriC E.coli's origin of Chromosome replication
  (given here for orientation only)
• gal E.coli's gene for galactose utilization
• peprophage ends (site of integration)
• cos joined sticky ends of vegetative DNA
  sometimes called ve ("vegetative ends")
• int gene for the enzyme integrase
• c gene for lambda repressor to maintain lysogeny
• Q another gene concerned with lysogeny
• h the last of the many capsomer genes

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Normal Excision lytic pathway
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Abnormal Excision
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??????????? ( lambda phage )

• Genetic engineering
• Specialized Transduction

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Cosmid vector

• A sequence marked "ori" for DNA replication in
  bacteria
• Ampr for ampicillin selection in bacteria
• A sequence marked MCS (multiple cloning site)
  that is a polylinker containing unique
  restriction sites and two phage RNA polymerase
  promoters (T7 and T3) in opposing directions.
• Two cos sequences, separated by an Xba1 site
• An origin of DNA replication from SV40 (permits
  replication and copy number amplification in many
  eukaryotic cells, in the presence of SV40 T
  antigen protein)
• Neor for selection in eukaryotic cells with the
  neomycin antibiotic analog G418

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Genetic engineering

• Cos site
• Cosmid vector
• ( 5 kbp)
• Large flagment
• ( 35-45 kbp)
• In vitro packaging
• Select for drug resistant

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• Insertional vector
• Replacement vector

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Transduction Viruses are the vector as they
move genes from cell to cell.

• Specialized Transduction
• Generalized Transduction

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Generalized Transduction A piece of host DNA
gets packaged by mistake, instead of the phage
DNA.  This rare event results in a phage
delivering only bacterial DNA to the next host. 
The DNA then recombines homologously, replacing
the host allele.
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Specialized Transduction in which a lysogenic
prophage recombines itself out of the genome (by
site-specific recombination) and mistakenly
includes a piece of bacterial DNA.  The resulting
phage progeny can infect cells to produce
lysogens with a second copy of the allele they
had packaged, attached to the phage DNA.
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