Difference between revisions of "Part:BBa K4632002:Design"
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C.What we have done?（SCAU-China 2023）
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In our design, we aimed to introduce a gene fragment encoding an active Cry3A-like toxin (66.6 kDa) into Escherichia coli using the pET30a vector to confer it with the ability to produce an active Cry3A-like toxin.
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To achieve secretion expression, we added a signal peptide sequence, OmpA, to the N-terminus of Cry3A-like toxin. This was done to direct the transport of Cry3A-like toxin to the extracellular space. OmpA is a well-established signal peptide in Escherichia coli for the secretion expression of foreign proteins. Our SDS-PAGE results confirmed the successful secretion expression of Cry3A-like toxin.
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Furthermore, we fused a 6×His tag to the C-terminus of Cry3A-like toxin to facilitate subsequent protein purification and Western blot-specific characterization experiments.
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In addition, to validate the toxicity of the designed Cry3A-like toxin against red imported fire ants, we selected homologous receptors of Cry3A-like toxin known from NCBI in red imported fire ants. We then conducted molecular docking studies to assess the protein-protein interaction capability of Cry3A-like toxin, thus evaluating its toxic effects. Detailed results are presented in the characterization section.
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Revision as of 17:32, 8 October 2023


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Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 1813
    Illegal PstI site found at 295
    Illegal PstI site found at 304
    Illegal PstI site found at 1456
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 1813
    Illegal PstI site found at 295
    Illegal PstI site found at 304
    Illegal PstI site found at 1456
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 1813
    Illegal BglII site found at 533
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 1813
    Illegal PstI site found at 295
    Illegal PstI site found at 304
    Illegal PstI site found at 1456
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 1813
    Illegal PstI site found at 295
    Illegal PstI site found at 304
    Illegal PstI site found at 1456
    Illegal AgeI site found at 1495
  • 1000
    COMPATIBLE WITH RFC[1000]