Part:BBa_K3221200
Galactose-mediated induction expression system for S.cerevisiae, yeGFP as a reporter
GAL system is a galactose-mediated induction system in yeast. To be specific, GAL1 promoter can induce downstream gene expression under the control of GAL4 protein. What’s more, in order to simplify the verification of GAL1 promoter function, we chose the yeGFP as a reporter instead of Cns1 and Cns2 in our project.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 150
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1199
Design: In order to demonstrate the function of GAL1 promoter, we cultured transformed yeast and WT in YPD medium and YPG medium separately. Through detecting green fluorescence both in Fluorescence microscope(Figure.1 (a)) and plate reader(Figure.1 (b)) to verify the function of GAL1 promoter.
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Figure.1 (a) Fluorescence observation of S.cerevisiae BY4741 transformed pYES2-yeGFP. Series 1: BY4741 WT in YPD mediuM, Series 2: BY4741 WT in YPG medium, Series 3: BY4741 transformed BY4741 in YPD medium, Series 4: Transformed BY4741 in YPG medium. Visible Fluorescence in Series 4. (b) The fluorescence measurement of S.cerevisiaeBY4741 transformed pYES2-yeGFP. YPD: YPD medium, YPG: YPG medium, BY4741 WT: S.cerevisiae, BY4741 wildtype, BY4741 transformed: S.cerevisiae, BY4741 transformed plasmid pYES2-yeGFP, excitation light is 488nm and emission light is 525nm.
Result & conclusion: Generally, we verified the function of GAL1 promoter by observing green fluorescence in B4(a) and B6(a). In other words, the GAL1 promoter could induce expression of downstream gene (yeGFP in this experiment) in galactose.
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