Part:BBa_K1642010
Expression of Halorhodopsin under the control of PcpcG2
PcpcG2-HR is a plasmid used to test whether the protein halorhodopsin works as we expect.
Up and Down are sequences used for homologous recombination.
PcpcG2 is the promoter of cpcG2. CcaS phosphorylates CcaR under green light, and phosphorylated CcaR binds to promoter of cpcG2 to induce expression of cpcG2.
RBS is the RNA binding site, which will hopefully increase the expression level of HR.
Halorhodopsin (HR) proteins are light-driven inward-directed chloride pumps from halobacteria. We use this as our biodesalination driver which confers cyanobacteria the ability to absorb chloride to a significant degree.
Terminator is the transcription terminator for the E.coli RNA polymerase.
PcpcB is a constitutive promoter in cyanobacteria, and Kana is gene sequence for kanamycin resistance, so that bacteria which are successfully transformed this plasmid can be screened on the plate.
Based on PcpcG2 and halorhodopsin, we established a biodesalination system which relies on red and green light. The light colors in the three stages are red light, green light and white light respectively.
In the growth stage, red light is enough for photosynthesis of cyanobacteria, which has been proved to be feasible by a recent report(Abe, K. et al. 2014). To further confirm it, we determined the growth curve under red light. This growth rate is no slower than that of the stain under natural light as long as we adjust the light intensity.
In the induction stage, the green light induces the expression of HR. Additionally, green light can’t provide energy source for chlorophyll, thus creating a starvation condition. And the starvation can inhibit active export of sodium, which is essential for biodesalination in the working stage.
In the working stage, halorhodopsin absorbs light to drive chloride import. Addtionally, cyanobacteria will regain energy under natural light. The biodesalination process controlled by PcpcG2 is shown in Figure 1.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 3398
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 481
Illegal NgoMIV site found at 966
Illegal NgoMIV site found at 1125
Illegal NgoMIV site found at 1446
Illegal AgeI site found at 1317 - 1000COMPATIBLE WITH RFC[1000]
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