Part:BBa_K2627007
Light-induced crRNA targeting gltA
This part consists of CcaS-CcaR system light-induced promotor PcpcG2-172 and a small guide RNA(crRNA) from Type I-E CRISPRi system. This can be induced upon green light to express crRNA targeting gltA gene to inhibit TCA cycle.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Results
After the verification of the culture condition, we investigated the dynamic regulation effects of targeting gltA with light-controlled system through switching red light to green light at 2h, 5h and 9h. As shown in figure , all strains induced at different time grow better compared with illuminating at 0h. Bacteria growth was inhibited the most when being induced at 5h, showing an obvious palliation after sensing green light. The strains induced at the early-log phase (2h) grew slower than the control (upon red illumination), whereas it continuously grew and maintained a relatively high K value. It was supposed that cells growing at a moderate speed at early consumed less nutrition, thus drove the biomass constantly increase and had a higher level of K value. In accordance with the hypothesis, we concluded the negative control without crRNA targeting gltA which presented a lowest level of K value consumed too much nutrition so that it can’t satisfy the need of cell in late period of growth. In addition, inducing crRNA expression at 9h through green light, we found there were little growth variance between the strains green light induced at 9h and the control illuminated at red light continuously. It was supposed that at the stationary phase there was nearly no effect for inhibiting TCA cycle to repress the cell’s growth, which might due to its metabolic flow. The difference between the regulation effect of different induction time isn’t obvious through light switching compared with L-arabinose, might due to light sensor CcaS/CcaR system which only have 4.8-fold difference between red/green light induction.
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