Part:BBa_K4156097

pCadC-Bxb1

pCadC-Bxb1 is constructed with pH-sensitive promoter pCadC and serine integrase Bxb1.

Usage and Biology

pCadC is regulated by the membrane lineage activator protein (CadC) and exhibits higher activity in acidic medium than in neutral pH medium. It is used in response to acidic conditions to allow the strain to rapidly localize to tumor cells and activate downstream gene transcription.Bxb1 is used to control downstream logic gate switching and to provide gene signaling amplification.

In vitro characterization and data analysis of the reported strains

To improve signaling stability as well as accuracy, we added Amplifying genetic switches based on serine integrase (Bxb1,TP901) to the R reporter(BBa_) to construct the AR reporter. Fig 1 indicates pH (pCadc) induced AR reporters with homogenized fluorescence intensity (mRFP/Cell). In contrast to Fig 1 and 2, the fluorescence intensity of the AR reporter appeared more stable over time at pH 7.3 and was higher than that of the R reporter at pH 5.8, 6.3, and 7.3. This result indicates that the addition of amplifying genetic switch enhances the reporter intensity and robustness of the lactate biosensor.

Figure 1: Induction of downstream gene mRFP expression over time by the AR reporter consisting of pCadC+Switch+mRFP at different pH values.

Figure 2: Induction of downstream gene mRFP expression over time by the AR reporter consisting of pCadC +mRFP at different pH values.

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 368
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 642
  Illegal XhoI site found at 729
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1476