Coding

Part:BBa_K4081888

Designed by: Mengfei Guo   Group: iGEM21_Jianhua   (2021-10-15)


CDT2


Biology and Usage

The N. crassa cellobiose transporter, CDT2, is likely to be a good platform on which to start protein engineering to improve 2′FL transport. CDT2 has already been shown to be tolerant of mutation by directed evolution experiments that improved its ability to transport cellobiose.


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
    COMPATIBLE WITH RFC[1000]


Design and Properties

We use CDT2 from Neurospora crassa to export 2′FL across the cell membrane. The result of SDS-PAGE showed that we successfully express CDT2 in BY4741(figure1）.

Figure 1. The result of SDS-PAGE. Line: Marker; line 2: Express CDT2(59kD); line 3: Control.

Experimental approach

Transform CDT2 into BY4741 by lithium acetate conversion method to integrate genes into the genome of BY4741 for expression. Screen for transformants by AbA-YPD selection medium. Extract yeast total protein. Use SDS-PAGE to test whether the CDT2 is successfully expressed.



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