Part:BBa_K5159026
pCIT1-GFP-LIP2t
This part is constructed to test the strength and sensitivity to ammonium concentration of the inducible promoter pCIT1 in Yarrowia lipolytica. The 1500 bp native pCIT is induced by nitrogen starvation, making it a suitable candidate for implementing the stage control strategy by turning on the PUFA synthase expression under this lipid accumulation-favoring condition. However the promoter, especially its inducibility, is not well characterized.
The fluorescence intensity serves as an indicator of the expression level and thus the promoter strength under different inducing conditions. The fluorescence intensity measured was compared with that from pTEFin-GFP-LIP2t expression in Yarrowia lipolytica to obtain a relative value. To ensure the comparability, identical spacers and terminator settings as the pTEF-GFP-LIP2t cassette are applied.
During testing, the average fluorescence was measured, blanked with untransformed groups and normalized with OD600 to minimize influence from other factors and the differences between individual cells transformed.
The reporter gene in the construct can be changed to hrGFP, luciferase, eGFP or other fluorescent proteins to achieve a similar goal, each having its own advantage.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal XbaI site found at 738
Illegal XbaI site found at 1462 - 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 2361
Illegal XhoI site found at 796
Illegal XhoI site found at 925 - 23INCOMPATIBLE WITH RFC[23]Illegal XbaI site found at 738
Illegal XbaI site found at 1462 - 25INCOMPATIBLE WITH RFC[25]Illegal XbaI site found at 738
Illegal XbaI site found at 1462
Illegal AgeI site found at 1023 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 2144
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