Part:BBa_K316007

Recombinant GFP-XylE protein under Pveg promoter

This construct contains BBa_K316006 under the control of Pveg promoter BBa_K143053. It is designed so that the XylE activity is substantially reduced untill such a time when a TEV protease is added to the system and transcribed. TEV protease cleavable linker is positioned between the two proteins. Once the linker is cleaved, XylE is free to tetramerise and assume full activity. GFP is His tagged at the 5' end to facilitate purificaiton for in-vitro assays. This construct does not have a terminator at the end, another construct BBa_K143008 contains a double terminator BBa_B0014.

For more information about XylE, it's substrate and spectrophotometric assays, please see BBa_K316003 or our wiki[http://2010.igem.org/Team:Imperial_College_London/Results]

Safety

The substrate XylE works on is a chemical called catechol. It is classed as irritant in the EU but as toxic in the USA, as well as being a possible carcinogen. It should therefore be handled with care and proper safety equipment. More information is available on the Material Safety Data Sheet[http://www.sciencelab.com/msds.php?msdsId=9927131].

Structure and Features

Figure I. Graphical representation of the GFP-XylE construct with associated Pveg promoter, tags and linkers.

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 NgoMIV site found at 1229
  Illegal NgoMIV site found at 1401
  Illegal AgeI site found at 1752
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 785
  

For more information about our project please visit our wiki [http://2010.igem.org/Team:Imperial_College_London].