Part:BBa_K2824005

lldPRD operon promoter-Luxs

Long Description

The purpose of the construction of this part is to modify the quorum sensing of E. coli. LuxS gene can produce LuxS protein and catalyze the formation of AI-2., a signaling molecule of the quorum sensing pathway.. When the promoter of LuxS gene is replaced by LLDPRD operon promoter, the LuxS gene will be activated in the presence of lactic acid, thus the quorum-sensing can be started.

Characterization

The part can also be proved to work well in experiments: when there is no lactic acid, the LuxS gene is barely expressed, with very little background level of gene expression. When lactic acid exists, the LuxS gene is expressed and the quorum sensing of Escherichia coli is activated, so that GFP is produced (Figure 1). By controlling the concentration of lactic acid to detect the fluorescence intensity of different GFP, the following results can be obtained: When the concentration of lactic acid was 2mM, the fluorescence intensity of the bacteria co-expressed with lldPRD operon promoter-LuxS plasmid and LsrA promoter-GFP reached the peak (Figure 2).

Figure 1: Characterization of LuxS under different concentrations of lactic acid and IPTG. The LuxS protein had been an obvious protein band between the last two marker bands of 15kDa and 25kDa. The molecular weight of the LuxS protein is about 17kDa. Lane 1, IPTG 0mM, lactic acid 0mM, O-LuxS; Lane 2, IPTG 0.5mM, lactic acid 0mM, O-LuxS; Lane 3, IPTG 1mM, lactic acid 0mM, O-LuxS; Lane 4, IPTG 0mM, lactic acid 2mM, O-LuxS; Lane 5, IPTG 0.5mM, lactic acid 2mM, O-LuxS; Lane 6, IPTG 1mM, lactic acid 2mM, O-LuxS; Lane 7, control( IPTG 0mM, lactic acid 0mM); Lane 8, IPTG 0mM, lactic acid 0mM, O-LuxS-lldR; Lane 9, IPTG 0.5mM, lactic acid 0mM, O-LuxS-lldR; Lane 10, IPTG 1mM, lactic acid 0mM, O-LuxS-lldR; Lane 11, IPTG 0mM, lactic acid 2mM, O-LuxS-lldR; Lane 12, IPTG 0.5mM, lactic acid 2mM, O-LuxS-lldR; Lane 13, IPTG 1mM, lactic acid 2mM, O-LuxS-lldR.

Figure 2:lldPRD operon promoter-LuxS work with LsrA promoter-GFP

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 505
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 603
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 335