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Regulatory

Part:BBa_K136004:Design

Designed by: Cyprien Maisonnier   Group: iGEM08_Paris   (2008-08-12)

Promoter of flhDC gene from E. coli flagella


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

The PCR on colony to amplify the promoter of flhDC did not work. We amplified both the promoter and the gene and then we did a specific amplification of the promoter from the full operon. We had a PCR amplicon of the right length this time.

The primers used to amplify the promoter and the gene are : 

  * Forward :

GTTTCTTCGAATTCGCGGCCGCTTCTAGAGTTGTATGTGCGTGTAGTGACGAGTACAG 

  * Reverse :

GTTTCTTCCTGCAGCGGCCGCTACTAGTATTATTAAACAGCCTGTACTCTCTGTTCATCC

Then to amplify only the promoter we used : 

  * Forward :

GTTTCTTCGAATTCGCGGCCGCTTCTAGAGTTGTATGTGCGTGTAGTGACGAGTACAG 

  * Reverse :

GTTTCTTCCTGCAGCGGCCGCTACTAGTATCCCACCCAGAATAACCAACTTTAT


Source

This part comes from the genome of E.coli K12 strain MG1655.

References

[1] Kalir S, McClure J, Pabbaraju K, Southward C, Ronen M, Leibler S, Surette MG, Alon U. Ordering genes in a flagella pathway by analysis of expression kinetics from living bacteria. Science. 2001 Jun 15;292(5524):2080-3.

[2] Shiraz Kalir, Uri Alon. Using quantitative blueprint to reprogram the dynamics of the flagella network. Cell, June 11, 2004, Vol.117, 713-720

[3] Kalir S, Mangan S, Alon U. A coherent feed-forward loop with a SUM input function prolongs flagella expression in Escherichia coli. Mol Syst Biol. 2005;1:2005.0006. Epub 2005 Mar 29.