Part:BBa_T9002

GFP Producer Controlled by 3OC6HSL Receiver Device

The luxR based receiver, F2620 (formerly I13270), controls the production of GFP. The GFP protein generator is the same as that found in I7101.

Characterization and improvement of BBa_T9002

(Characterized by Peking 2019)

We used this part to characterize the quorum sensing system. Induced by different concentration of AHL, the GFP expression level showed obvious positive relationship with AHL. The result and protocols are as follows.

Figure 1.The response curve of plux.

Experiment procedure:
1.E. coli DH5α transferred with PSB1C3-T9002 were incubated in LB liquid medium at 37℃, 220rpm overnight
2.Grown culture was 100-fold diluted in M9 medium for the same condition for 3 hours.
3.Grown culture was 40-fold diluted in M9 medium containing different concentration of AHL.
4.200 µl aliquots of each of the cultures were transferred into a flat-bottomed 96 well plate.
5.FI and OD600 was measured for 10 hours in microplate reader.

We used BBa_T9002 to characterize quorum sensing system. But we found that this part has an obvious homologous recombination. So we changed the terminator of GFP to avoid it.

Figure2 The design of quorum sensing system.

Figure3 A time-scale quorum sensing system can be transformed to spatial-level through a donor/receiver system. The donor cells, which merely express and release AHL, would activate the GFP expression of receiver cells through AHL diffusion. This is validated on the agar plate, by dropping the donor cells in the center and receiver cells around them with different distances. We found a progressive decrease in fluorescence as the receiver cells locate farther from central AHL donor.

Experiment procedure:
1.Incubate donor and receiver cells in LB medium overnight.
2.Grown culture was 100-fold diluted in M9 medium.
3.5 μL donor cell was dropped in the LB solid medium.
4.5 μL receptor cell was dropped in different distances in the same LB solid medium.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1004
  Illegal BsaI.rc site found at 1732