Composite

Part:BBa_K2542014

Designed by: Yaoqiang Zheng   Group: iGEM18_NEFU_China   (2018-10-05)


pFUS1-EGFP-tCYC

This plasmid allows a type yeast to sense the alpha factor secreted by alpha-type yeast and express green fluorescent protein. We modified the part(BBa_K775004) of the previous team to make the expression efficiency weaker,and make it more appropriate for suicide gene(Information Destroyed ).

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]

Experiment and Result

iGEM18_NEFU_China creat a new parts BBa_K2542014,which is based on BBa_K775004. BBa_K2542014 is more suitable for somewhere which needs low expression in yeast.It's allows a type yeast to sense the α factor secreted by α-type yeast and express green fluorescent protein. We modified the part BBa_K775004 of the previous team by turning mutating the sequence “AAAAAA” into “GCCACC” to make the expression efficiency weaker,and make it more appropriate for suicide gene(Information Destroyed ). We constructed two plasmids with different kozak sequences and transferred them into yeast. Then, we use SD-Trp medium to culture yeast, and add with the solution which containing α factor. The solution containing α factor is obtain by extracting the liquid supernatant of the culture medium for culturing α-type yeast. After we add 200ul the α factor sloution to each one and culture yeast for 20 hours,we measured the expression of GFP in yeast by flow cytometry.

T--NEFU China--Kozak sequence change result.jpg


[edit]
Categories
Parameters
None