IGEM:Caltech/2008/Project/ROS

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iGEM 2008

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Conjugation

  • Can get conjugation from Salmonella to E. coli [1]
    • Papers seem to assume that conjugation is quick, and maintenance is the hard part (unless we engineer a shuttle vector). Doesn't address specificity, though.
  • "Conjugation of S. typhi with E. coli F+ carrying P1CM+ gave three types of S. typhi CMr clones: those which carry the whole P1CMphage, those with the PldCM element, and those with nontransferable CMr."[2]
  • Can get transfer of an F' plasmid from E. coli to Salmonella[3]
  • Doug Tischer 21:12, 26 May 2008 (EDT)There exists a plasmid (pAT187) that can be transfered from E. coli to a wide range of gram-positive bacteria [4]
  • Doug Tischer 17:42, 27 May 2008 (EDT)Surface exclusion seems to be the name given to the process that prevents a bacteria containing a plasmid from conjugating and receiving the same plasmid again. In F plasmid, the are the traS and traT genes. traT inhibits the formation of mating aggregates. [5]
  • Doug Tischer 17:51, 27 May 2008 (EDT) Biobrick part BBa_J01000 (TraJf) is a transcriptional activator that turns on the expression of sex pili forming genes off of the F plasmid. This could be used to trigger the oxidative burst upon conjugation instead. See this page for information explaining this and other conjugation related Biobricks.

Quorum Sensing

  • Given the complexity of a conjugation based triggering system, perhaps quorum sensing would be more feasible. Interspecies signaling is carried out through acyl-homoserine lactone (AHL). These are very specific, and each type of bacteria produces there own unique one. It only takes one cytoplasmic receptor protein to bind to the AHL and activate transcription. The production of AHL is associated with a switch from a benign to a pathogenic form in several bacteria. We could make our cells target Salmonella by giving them the receptor for the Salmonella AHL. The only thing we would need to change is to make the detection quite sensitive, so that our cells kill the Salmonella become pathogenic. [6]
  • lsrB [7]

Specificity

  • Could we detect transmission of the Salmonella virulence plasmid?
    • "The second experiment was to physically observe the transferred virulence plasmid in the recipient strain.... Therefore, E. coli was chosen to be the recipient in matings with Salmonella.... Transconjugants were obtained at a frequency of approximately 2 × 10−5 transconjugants/donor.... Ethidium bromide staining of the pulsed-field gel shows that the restriction pattern of the transconjugant genomes matches that of the E. coli recipient, except for the acquisition of the virulence plasmid from the Salmonella donor. The identity of the virulence plasmid was confirmed by subsequent Southern hybridization with the kanamycin resistance gene of MudJ.... This is clear evidence that conjugation has occurred."[8]
    • sdiA transcription factor? [9]
    • Or spvR[10, 11]. It functions in E. coli and is necessary for virulence. So our bacteria are floating around waiting for conjugation from a pathogenic Salmonella. Once that happens, a spvR promoter turns on, triggering the ROS. The low conjugation frequency is troubling. However, some of that might be low efficiency genomic integration.

H2O2 Transport

  • Diffusion of H2O2 across E. coli membranes is "exhibit substantial, but limited, permeability to H2O2." The review suggested some aquaporins in yeast increase cell membrane permeability to H2O2. [12]
  • The strain JI377 (a deletion for katE, katG, and ahp) cannot scavange H2O2 whatsoever and excretes it into the medium. [13]

References

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  1. Error fetching PMID 12843024: [makanera]
  2. Error fetching PMID 5327907: [kondo]
  3. Error fetching PMID 7009564: [lenny]
  4. Error fetching PMID 2656662: [Mazodier]
  5. Error fetching PMID 357413: [Achtman]
  6. Error fetching PMID 14597753: [Federle]
  7. Error fetching PMID 15350213: [miller]
  8. Error fetching PMID 9973370: [ahmer]
  9. Error fetching PMID 9495757: [ahmer2]
  10. Error fetching PMID 9286993: [grob]
  11. Error fetching PMID 9421907: [wilson]
  12. Error fetching PMID 16566894: [Bienert]
  13. Error fetching PMID 11717276: [Seaver]
  14. Error fetching PMID 12496934: [Plainkum]
  15. Error fetching PMID 11698678: [Firbank]
  16. Error fetching PMID 11707617: [Sun]
  17. Error fetching PMID 11317240: [Uehara]
  18. Error fetching PMID 9490070: [Toomey]
  19. Error fetching PMID 15581579: [Whittaker]
All Medline abstracts: PubMed HubMed
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