Part:BBa_K3600008

prYCF1-yomScarlet-I-tADH1

This cassette was assembled from 6 part plasmids via Golden Gate assembly using the BsmbBI restriction enzyme. The two connectors (pYTK002 and pYTK072), as well as the terminator (pYTK053) and the Ampicilin resistance/backbone (pYTK095) part plasmids are part of the YTK toolkit developed by Lee et al.¹ and can be ordered online. The promoter part plasmid (BBa_K3600002) and the reporter part plasmid (BBa_K3600010) were synthesized by us.

Usage and Biology

This cassette contains the YCF1 promoter upstream of a reporter gene (mScarlet-i). YCF1 is a stress response gene in s.cerevisiae that is regulated by the YAP1 master regulator². This cassette is then assembled with a part plasmid containing Lys2 homology sequences (BBa_K36000012) in another Golden Gate BsmBI assembly, to give a final integration cassette (BBa_K3600015).

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 935
  Illegal EcoRI site found at 1665
  Illegal XbaI site found at 944
  Illegal XbaI site found at 1842
  Illegal SpeI site found at 2810
  Illegal PstI site found at 2819
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 935
  Illegal EcoRI site found at 1665
  Illegal SpeI site found at 2810
  Illegal PstI site found at 2819
  Illegal NotI site found at 923
  Illegal NotI site found at 2829
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal suffix found in sequence at 2412
  Illegal EcoRI site found at 935
  Illegal EcoRI site found at 1665
  Illegal BglII site found at 1280
  Illegal BglII site found at 2283
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 935
  Illegal EcoRI site found at 1665
  Illegal XbaI site found at 944
  Illegal XbaI site found at 1842
  Illegal SpeI site found at 2810
  Illegal PstI site found at 2819
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 935
  Illegal EcoRI site found at 1665
  Illegal XbaI site found at 944
  Illegal XbaI site found at 1842
  Illegal SpeI site found at 2810
  Illegal PstI site found at 2819
  Illegal NgoMIV site found at 2767
  
• 1000
  COMPATIBLE WITH RFC[1000]

References

[1] Lee, M. E., DeLoache, W. C., Cervantes, B. & Dueber, J. E. A Highly Characterized Yeast Toolkit for Modular, Multipart Assembly. ACS Synth. Biol. 4, 975–986 (2015).

[2] Gomes DS, Fragoso LC, Riger CJ, Panek AD, Eleutherio EC. Regulation of cadmium uptake by Saccharomyces cerevisiae. Biochimica et Biophysica Acta (BBA)-General Subjects. 2002 Oct 10;1573(1):21-5.