Part:BBa_K1679025

pT7-4 +RiboJ + GFP + LVA + 3 terminnators

This device is design for protein expression and promoter measurement. It constructed with a T7 RNA polymerase which has other 3 mutations, a RiboJ, a green fluorescent protein, a LVA degradation tag, a T7 terminator, a rrnB T1 terminator and a rrnB T2 terminator. The four T7 RNA polymerases are a family with different activity, the T7 RNA polymerase in this device have approximate relative activity of 4%.

Usage and Biology

Fig.1 T7 protein expression device.

Fig.2 The 4 T7 RNA polymerase and their approximate relative activity.

Experiment

We expressed GFP on this device in BL21(DE3). Three monoclonal colonies were selected and culture until OD600 0.5(37℃, 220rpm).We calculated the Fluorescence(excitation wavelength-584 nm and emission wavelength-607 nm) and OD600.

Fig.3 Three colonies with this part and three colonies withBBa_K1679026 were selected to measure.

Fig.4 The fluorescent/OD of three colonies in fig.3 were averaged.

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
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 1019
  
• 1000
  COMPATIBLE WITH RFC[1000]