Generator

Part:BBa_K1741004

Designed by: Marta Żardecka   Group: iGEM15_UAM_Poznan   (2015-09-17)
Revision as of 07:27, 27 September 2015 by MartaZ (Talk | contribs)

sfGFP under melibiose promoter (improved 5'UTR)

Twice as high expression (comparing to BBa_K1741003) has been obtained by 5’UTR editing – removal of potential secondary structure which involved RBS and a better positioning of RBS – 7 nt upstream AUG start codon. The modified 5’UTR results in about twice higher expression upon melibiose addition to the M9 or 2xLB medium. The promoter is still weaker than comercial araBAD or rhaBAD promoters, so can be the promoter of choice if lower expression level is necessary for efficient folding or secretion of a recombinant protein.

Design

Legend:

MelS [BBa_K1741004]

UAMpoznanmelS.jpg

MelWT [BBa_K1741003]

UAMpoznanmelWT.jpg

Results

Legend:

MelS [BBa_K1741004]
MelWT [BBa_K1741003]

Teamuampoznanmelibiosem9graph.png

Teamuampoznanmelibioselbgraph.png

The sfGFP fluorescence was measured using Tecan fluorometer.

We also checked the tightness of our promoters.

All of our promoters are induced only by their respective sugars with a small exception of xylose promoters being slightly induced by arabinose and arabinose promoters being slightly induced by xylose. All of our promoters are repressed by glucose. Teamuampoznanpromoterscomparisongrpah.png


Sequence and Features


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
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 207


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Parameters
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