Coding

Part:BBa_K2796028:Design

Designed by: Shuangshuang Pu   Group: iGEM18_LZU-CHINA   (2018-10-04)


Exosome booster

  • The part we designed and plasmid map are shown below:
    • 2018 LZU-CHINA experimental design1.png

    Agarose gel electrophoresis

    Three genes of exosome booster (hSDC4, Steap3, nadB) were detected by agarose gel electrophoresis. These three genes could help to enhance cell to secrete exosome.

    2018 LZU-CHINA experimental Gel-1.png

    Western Blotting

    Protein sample extracted from engineering HEK293T cell was divided into two parts.One was used for staining to verify the presence of protein.

    2018 LZU-CHINA experimental Gel-2.png

    The staining results indicated that the protein was extracted from engineering HEK293T cell. Another sample was used to do wester blotting to validate whether the exosome booster gene express or no.

    2018 LZU-CHINA experimental Gel-3.png

    The WB result showed that all three exosome booster genes were successfully expressed in HEK293T. Because nadB is derived from bacteria, there's no corresponding antibody. We added 6 histidine at the 3 'end of the nadB ‘s coding site and used histidine antibodies for its detection.




    Assembly Compatibility:
    • 10
      COMPATIBLE WITH RFC[10]
    • 12
      COMPATIBLE WITH RFC[12]
    • 21
      INCOMPATIBLE WITH RFC[21]
      Illegal BglII site found at 192
      Illegal BglII site found at 1777
    • 23
      COMPATIBLE WITH RFC[23]
    • 25
      INCOMPATIBLE WITH RFC[25]
      Illegal AgeI site found at 2657
      Illegal AgeI site found at 3277
    • 1000
      INCOMPATIBLE WITH RFC[1000]
      Illegal SapI site found at 1367

    Design Notes

    This part will help our cell to increase exosome secretion.


    Source

    NCBI

    ===References===