Composite

Part:BBa_K4759217

Designed by: Jianhua Chen   Group: iGEM23_Jiangnan-China   (2023-10-07)
Revision as of 16:21, 10 October 2023 by JianghuaChen (Talk | contribs)


T7-RBS1-Fdr-0978-RBS2-Fdx-1499-linker-GFP1-10

Generally, the method of determining whether the redox partner is suitable is through tedious steps such as the construction of plasmids, heterologous expression, construction of catalytic systems, and detection of conversion rate after catalysis. Therefore, we wanted to find a convenient way to do a quick screening. We used the fluorescent protein sfGFP to successfully construct a sensor to detect redox partners. sfGFP is a superfolder fluorescent protein that emits green light when irradiated with ultraviolet light. What is special about it is that it can be broken into two parts. We divide sfGFP into sfGFP-1-10 and sfGFP-11, and although these two parts are cut off, there is an interaction force between them, and as long as they are properly folded in space, they will emit light again. Thus, four iron redox proteins are fused to the N-terminus of sfGFP-1-10 and Olep to the C-terminus of sfGFP-11, respectively, to obtain the recombinant plasmid pRSFDuet-BM3-GFP-1-10-GFP-11-oleP, pRSFDuet-camA-camB-GFP-1-10-GFP-11-oleP, pRSFDuet-FdR_0978-Fdx_1499-GFP-1-10-GFP-11-oleP, pRSFDuet-petH-petF-GFP-1-10-GFP-11-oleP The above four recombinant plasmids are converted to BL21(DE3) to obtain recombinant strains G2 strain to G5 strain.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 1306
    Illegal PstI site found at 368
    Illegal PstI site found at 694
    Illegal PstI site found at 1009
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 1306
    Illegal PstI site found at 368
    Illegal PstI site found at 694
    Illegal PstI site found at 1009
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 1306
    Illegal BglII site found at 1124
    Illegal BglII site found at 2295
    Illegal BamHI site found at 1300
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 1306
    Illegal PstI site found at 368
    Illegal PstI site found at 694
    Illegal PstI site found at 1009
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 1306
    Illegal PstI site found at 368
    Illegal PstI site found at 694
    Illegal PstI site found at 1009
    Illegal NgoMIV site found at 712
    Illegal AgeI site found at 784
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 1429


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