Difference between revisions of "Part:BBa J45999:Design"
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===Design Notes===
 
===Design Notes===
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A key challenge in our work to reprogram bacterial odor was to ensure that the natural, fecal odor of ''E. coli'' did not overpower our engineered wintergreen and banana odors.  Indole was suggested to be the primary component of the fecal odor of ''E. coli'' (E Pichersky, personal communication, 2008).  We confirmed that indole was the primary odor produced by ''E. coli'' by smelling LB Lennox medium supplemented with indole at a concentration comparable to that of LB cultures of ''E. coli'' strain MG1655 (~300 &mu;M) <cite>Wang-2001</cite>.  In nature, ''E. coli'' uses indole for intercellular signaling in biofilm formation <cite>Martino2003</cite>.  Since indole is not essential for cell viability, we could reprogram bacterial odor by modifying cellular metabolism for decreased indole production.
  
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* CGSC#: 7602
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Indole is a product of the tryptophanase enzyme encoded by the ''tnaA'' gene of the ''tna'' operon in ''E. coli'' <cite>Snell-1975</cite>.  Mutations to the ''tna'' operon can reduce indole levels <cite>Rezwan-2004</cite>. We tested four ''E. coli'' strains as potential odor-free chassis for our engineered system: YYC912, JC12337, MEB61, and MB408 (CGSC 7602, CGSC 6373, CGSC 6836, and CGSC 7152, respectively) <cite>Chang-1994, Ream-1980, Edwards-1982</cite>. The four strains all carry mutations in the ''tnaA'' gene. By smelling overnight liquid LB cultures of each strain, we selected ''E. coli'' strain YYC912 as an odor free chassis for our engineered system. We confirmed via gas chromatography analysis that ''E. coli'' strain YYC912 produced no measurable indole in comparison to ''E. coli'' strain TOP10.
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* Strain Designation
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**Designation: YYC912
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**Source Person: Cronan & Chang
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**Choice: 1 
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* Sex(Hfr,F+,F-,or F'): F-
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* Mutations
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**DE(argF-lac)169  
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**LAM-
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**poxB15::lacZ::CmR 
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**IN(rrnD-rrnE)1 
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**rph-1 
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**tnaA5 
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* No. of Muts Carried: 6
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* Special Uses: lac fusion-stat.phase promoter
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* Has_Partner_Isogenic_WRT: poxB15::lacZ::CmR
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* Isogenic_Partner_Is: ZK126
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* References
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**Chang, Y.-Y., A.Y. Wang, J.E. Cronan 1994. Expression of Escherichia coli pyruvate oxidase (PoxB) depends on the sigma factor encoded by the rpoS(katF) gene. Mol.Microbiol. 11:1019-1028 
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* ID#: 64826
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===Source===
 
===Source===
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We thank Mary Berlyn for useful advice regarding ''tna'' operon mutants and [http://cgsc.biology.yale.edu/ The Coli Genetic Stock Center] for ''E. coli'' strains [http://cgsc2.biology.yale.edu/Strain.php?ID=64826 YYC912 (CGSC 7602)], JC123337 (CGSC 6373), MEB61 (CGSC 6836), and MB408 (CGSC 7152).
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This strain was obtained from the E.coli Genetic Resources: CGSC, The Coli Genetic Stock Center http://cgsc.biology.yale.edu/.  Strain YYC912.
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===References===
 
===References===
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<biblio>
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#Wang-2001 pmid=11418561
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#Snell-1975 pmid=236639
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#Rezwan-2004 pmid=15489459
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#Chang-1994 pmid=8022274
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#Ream-1980 pmid=6255290
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#Edwards-1982 pmid=7052072
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#Martino-2003 pmid=14569285
 +
</biblio>
 
Chang, Y.-Y., A.Y. Wang, J.E. Cronan 1994. Expression of Escherichia coli pyruvate oxidase (PoxB) depends on the sigma factor encoded by the rpoS(katF) gene. Mol.Microbiol. 11:1019-1028
 
Chang, Y.-Y., A.Y. Wang, J.E. Cronan 1994. Expression of Escherichia coli pyruvate oxidase (PoxB) depends on the sigma factor encoded by the rpoS(katF) gene. Mol.Microbiol. 11:1019-1028

Latest revision as of 09:50, 30 June 2014

E. coli strain YYC912; Odor-free chassis


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

A key challenge in our work to reprogram bacterial odor was to ensure that the natural, fecal odor of E. coli did not overpower our engineered wintergreen and banana odors. Indole was suggested to be the primary component of the fecal odor of E. coli (E Pichersky, personal communication, 2008). We confirmed that indole was the primary odor produced by E. coli by smelling LB Lennox medium supplemented with indole at a concentration comparable to that of LB cultures of E. coli strain MG1655 (~300 μM) Wang-2001. In nature, E. coli uses indole for intercellular signaling in biofilm formation Martino2003. Since indole is not essential for cell viability, we could reprogram bacterial odor by modifying cellular metabolism for decreased indole production.

Indole is a product of the tryptophanase enzyme encoded by the tnaA gene of the tna operon in E. coli Snell-1975. Mutations to the tna operon can reduce indole levels Rezwan-2004. We tested four E. coli strains as potential odor-free chassis for our engineered system: YYC912, JC12337, MEB61, and MB408 (CGSC 7602, CGSC 6373, CGSC 6836, and CGSC 7152, respectively) Chang-1994, Ream-1980, Edwards-1982. The four strains all carry mutations in the tnaA gene. By smelling overnight liquid LB cultures of each strain, we selected E. coli strain YYC912 as an odor free chassis for our engineered system. We confirmed via gas chromatography analysis that E. coli strain YYC912 produced no measurable indole in comparison to E. coli strain TOP10.

Source

We thank Mary Berlyn for useful advice regarding tna operon mutants and [http://cgsc.biology.yale.edu/ The Coli Genetic Stock Center] for E. coli strains [http://cgsc2.biology.yale.edu/Strain.php?ID=64826 YYC912 (CGSC 7602)], JC123337 (CGSC 6373), MEB61 (CGSC 6836), and MB408 (CGSC 7152).

References

<biblio>

  1. Wang-2001 pmid=11418561
  2. Snell-1975 pmid=236639
  3. Rezwan-2004 pmid=15489459
  4. Chang-1994 pmid=8022274
  5. Ream-1980 pmid=6255290
  6. Edwards-1982 pmid=7052072
  7. Martino-2003 pmid=14569285

</biblio> Chang, Y.-Y., A.Y. Wang, J.E. Cronan 1994. Expression of Escherichia coli pyruvate oxidase (PoxB) depends on the sigma factor encoded by the rpoS(katF) gene. Mol.Microbiol. 11:1019-1028