Part:BBa_I739001

Constitutive expression cassette for TetR +LVA (J23100.B0034.C0040.B0015)

Part Structure

The Biobrick encodes LVA-tagged TetR (BBa_C0040) under control of the constitutive promoter BBa_J23105 followed by the ribosome binding site BBa_B0034. The transcription of tetR is terminated by the double terminator BBa_B0015.

Mode of Action

TetR binds to the pTet regulator (BBa_R0040) and represses transcription. If the inducer [http://openwetware.org/wiki/ATc anhydrotetracycline (ATc)] is added, TetR action is inhibited and the promoter gets derepressed.

Purpose

This Biobrick was designed for the [http://2007.igem.org/ETHZ ETHZ iGEM 2007 project] and belongs to the constitutive part of the system. In the project description, this part is also termed Part 1. The constitutively synthesized TetR interacts with the double promoters BBa_I739102 and BBa_I739106 which are parts of composites BBa_I739015 and BBa_I739010 respectively. A similar (but not used) construct in this context is BBa_Q04400.

Testing

Checked for uniqueness of restriction enzyme cleavage sites:
Eco: ok
Xba: ok
Spe: ok
Pst: ok

Additional characterization of BBa I739001 by [http://2016.igem.org/Team:LMU-TUM_Munich Munich 2016]

BioBrick I739001 BBa I739001 was ligated upstream of BioBrick BBa_K2170221 (TetO-OmpA) which was then ligated upstream of BioBrick BBa_K2170142 (NanoLuc-Strep Tag-double terminator) to assemble BioBricks BBa_K2170141 (TetR-TetO-OmpA) and BBa_K2170053 (TetR-TetO-OmpA-NanoLuc-Strep Tag-double terminator. The functionality of both BioBricks could be confirmed by Nano-Glo assaywhich is why the functionality of I739001 BBa I739001 could be validated as well.

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
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]