Part:BBa_K4779000:Design
CMPG:A yeast copper-induced reporter pathway with prm1 promoter
- 10INCOMPATIBLE WITH RFC[10]Illegal XbaI site found at 279
Illegal XbaI site found at 517
Illegal SpeI site found at 343
Illegal SpeI site found at 523
Illegal PstI site found at 49 - 12INCOMPATIBLE WITH RFC[12]Illegal SpeI site found at 343
Illegal SpeI site found at 523
Illegal PstI site found at 49 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 55
Illegal BamHI site found at 510
Illegal XhoI site found at 8 - 23INCOMPATIBLE WITH RFC[23]Illegal XbaI site found at 279
Illegal XbaI site found at 517
Illegal SpeI site found at 343
Illegal SpeI site found at 523
Illegal PstI site found at 49 - 25INCOMPATIBLE WITH RFC[25]Illegal XbaI site found at 279
Illegal XbaI site found at 517
Illegal SpeI site found at 343
Illegal SpeI site found at 523
Illegal PstI site found at 49
Illegal NgoMIV site found at 36 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 2213
Illegal SapI site found at 529
Design Notes
When designing the sequence in detail, one of the key considerations was how to convert copper ion signals into information molecule signals. We achieved this signal conversion through the pCup1 and MFα2 elements, and amplified the signals through a cascade amplification system based on the MAPK pathway. We leveraged the high sensitivity of the MAPK pathway to pheromone to enhance the system's sensitivity to copper ion concentration signals. Here, we placed the pheromone-responsive promoter (in this case, pprm1) in-frame with a yeast enhanced green fluorescent protein, thereby generating a measurable fluorescence signal and facilitating the quantification of copper ion concentrations.
Source
We obtained the gene sequences of the basic parts from IGEM, such as the promoter of Cup1 (BBa_K945002), MFα2 (BBa_K110005), the promoter of prm1 (BBa_K1346004), green fluorescent protein (GFP) (BBa_K2176000), and the terminator CYC1 (BBa_K4278703), and sent them to GENEWIZ (Suzhou Genewiz Biotechnology Co., Ltd.) for synthesis. The new composite part (CMPG) (BBa_K4779000) is also synthesized.