Part:BBa_K256007

J23119:NorR

Generator of NorR

This construct constitutively produce NorR (BBa_K256004) to supplement the function of NO sensing promoter, pNO (BBa_K256000).

Usage and Biology

NorR is a DNA-binding transcriptional activator, also known as B2709, NorR, YgaA and transcriptional regulator of the anaerobic response to reactive nitrogen species.

In E.coli, transcription factor NorR is being endogenously expressed. Yet, to reduce competitive binding of NorR between the endogenous NorR-NorVW intergenic region (in E.coli) and pNO, large scale production of NorR is necessary. This is accomplished by introducing the constitutively ON NorR generator.

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 AgeI site found at 716
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI.rc site found at 491
  

Applications of BBa_K256007

NorR is a DNA-binding transcriptional activator also known as B2709, NorR, YgaA and transcriptional regulator of the anaerobic response to reactive nitrogen species.

BBa_K256007 constitutively produce NorR ( BBa_K256004) to supplement the function of NO sensing promoter, pNO ( BBa_K256000).

Proof of existence

The successful construct J23119-B0034-NorR-B0015 was ligated to pSB3K2 vector, and was later introduced into Invitrogen’s TOP10 cells. Transformed cells were grown for 16 hrs, at 3 oC, 250 rpm, before undergoing cell lysis for SDS PAGE.

The SDS PAGE result was reflected in the gel photo. Production of 55 kDa NorR protein had been confirmed by the result.

Fig.1 55 kDa NorR band has been circled in red.

Characterization via SDS PAGE

To characterize the production of NorR in different environmental temperatures for incubation, the transformed cells were incubated at 3 different environmental temperatures (i.e. 30, 37 and 42°C).

Fig.2 55 kDa NorR band has been circled in red. NorR production was showed to vary with the culturing temperature.

It has been observed that at elevated temperature, i.e. 42°C, production of NorR had decreased as shown by the diminished intensity (see lane '42'). The production of NorR at 30°C and 37°C were comparable.