Part:BBa_K3581019

BBa_K880005 part without hagfish intermediate filament subunits with TorA signal peptide

This composite part includes composite part K880005, His tag BBa_K3581002, TorA signal peptide BBa_K3581014 and terminator BBa_B0015. The aim of this part is to maximize protein expression through the use of strong promoters and RBS optimal for protein secretion. The His tag will be used for purifying the protein and TEV protease cut site can cleave out the tag once the protein is purified. The signal peptide is flanked by HindIII cut sites which will allow us to secrete the protein extracellularly, when we can also cut out the signal peptide using HindIII to secrete them as inclusion bodies to determine the best method with highest yield. The terminator is a double terminator which is commonly used for termination.

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
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 123
  Illegal BsaI.rc site found at 255
  

References: 1. England, N. (n.d.). TEV Protease | NEB. Retrieved October 13, 2020, from https://international.neb.com/products/p8112-tev-protease#Product%20Information

2. Wikipedia. (2020, October 14). TEV Protease. Wikipedia. Retrieved October 13, 2020, from https://en.wikipedia.org/wiki/TEV_protease

3. Palmer, I., & Wingfield, P. T. (2004). Preparation and extraction of insoluble (inclusion-body) proteins from Escherichia coli. Current protocols in protein science, Chapter 6, Unit–6.3. https://doi.org/10.1002/0471140864.ps0603s38

4. https://parts.igem.org/Part:BBa_K880005

5. https://parts.igem.org/Part:BBa_B0015