Part:BBa_K2556333

Light control system with eGFP

Under dark conditions, the phosphate group is transferred from Yf1 protein to FixJ protein, the phosphorylated FixJ protein activates the PfixK2 promoter and then activates the downstream gene expression. When induced by light, the phosphorylation of Fixj protein will be blocked, and the expression of genes regulated by PfixK2 promoters will be inhibited.

Characterize

To further verify the effectiveness of the system, the plasmids with the system were transferred into different hosts and cultured for more than 24 hours under blue light and dark light respectively. The fluorescence value of eGFP in the culture medium was measured.

Experimental Results

Detection of expression efficiency of primary light control system in different hosts

Figure 1. expression efficiency in DH5α

Figure 2. expression efficiency in BL21

Figure 3. expression efficiency in 4MG1655

Figure 4. expression efficiency in BW25113

The dusk-eGFP-pUC57 plasmid was transferred into the host of DH5α,BL21,4MG1655 and BW25113, cultured under dark or blue light for a period of time and the fluorescence value of eGFP was detected. After 24 hours, the fluorescence value of the experimental group in the dark culture showed an obvious increasing trend. There was no significant increase in fluorescence value in both the experimental group under blue light and the negative control group.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 605
  Illegal NgoMIV site found at 677
  Illegal NgoMIV site found at 767
  Illegal NgoMIV site found at 785
  Illegal NgoMIV site found at 1297
  Illegal NgoMIV site found at 1590
  Illegal NgoMIV site found at 1684
  Illegal AgeI site found at 319
  Illegal AgeI site found at 1465
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1354
  Illegal BsaI.rc site found at 218
  

References

[1]Wang G, Lu X, Zhu Y, et al. A light-controlled cell lysis system in bacteria.[J]. Journal of Industrial Microbiology & Biotechnology, 2018:1-4.
[2]Wu H, Wang Y, Wang Y, et al. Quantitatively relating gene expression to light intensity via the serial connection of blue light sensor and CRISPRi[J]. Acs Synthetic Biology, 2014, 3(12):979.
[3]Gardner L, Deiters A. Light-Controlled Synthetic Gene Circuits[J]. Current Opinion in Chemical Biology, 2012, 16(3-4):292-299.