We created seven kinds of tandem promoters constructed by many single promoters in iGEM kit.
Fig 1. Schematics of our biobrick construction
For constructing the tandem promoter, we have to choose whether the promoter pBad or the constitutive promoter should be put in the former. Hence, we constructed two kinds of tandem promoters with pBad. One is the J23106 ahead of pBad, and the other is the J23106 behind pBad.
Fig 2. The RFP intensity of Pre C1 and Pre C2 after 24 hrs cultivation
Pre C1: pBad-J23106 Pre C2: J23106-pBad
When testing the RFP result, we invited our collaboration partner, TAS iGEM team, for help in the measurement. We found that Pre C2 is hardly induced by L-arabinose and the endogenous expression rate of Pre C2 is very low. On the other hand, the expression rate of Pre C1 is great when L-arabinose is added. The result showed that if we put pBad in the former, the expression of tandem promoter will be more stable. For more information about our collaboration, please visit our collaboration
After finding out that putting pBad in the former is the better choice, we generated five other tandem promoters with Strategy 3 to construct them rapidly. In these 5 tandem promoters, we selected five other constitutive promoters (BBa_J23102, BBa_J23118, BBa_J23106, BBa_J23110, BBa_J23113) in the same family of J23106. (Promoter strength：BBa_J23102＞BBa_J23118＞BBa_J23106＞BBa_J23110> BBa_J23116>BBa_J23113). We checked the strength of the tandem promoters by testing the RFP expression from each tandem promoter.
Fig 3. The RFP intensity of tandem promoters
According to the figures, when the concentration of L-arabinose is 0 M, the order of the expression rate of these tandem promoters is the same as that of the constitutive promoter individually. Besides, when the concentration of L-arabinose rises from 0 M to 0.025 M, the expression of RFP under the regulation of pJ102, pJ118, and pJ110 decreases while others rise. Interestingly, among these six tandem promoters, only the Pre C1 will be induced by L-arabinose significantly.