Part:BBa_K3271006

Gal4::His3-EXSP pSB1K3

The prefix and suffix (EXSP) of the pSB1K3 backbone are flanked by the upstream and downstream homology sequences of the Gal4 locus from S. cerevisiae, respectively. Between the upstream homology and the prefix is the His3 auxotrophic selection marker. To target a DNA part of interest to the Gal4 locus in S. cerevisiae, insert the DNA part within the prefix and the suffix using any method of your choice. Then, digest the plasmid using PmeI. This creates a DNA fragment comprised of the DNA part of interest flanked by the Ade4 locus upstream homology along with His3 auxotrophic selection marker and the downstream homology at the 5' and 3' ends of the part, respectively. Transform the digest into S. cerevisiae using a lithium acetate protocol of choice and plate on SC His-.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal prefix found at 3304
  Illegal suffix found at 3326
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 3304
  Illegal NheI site found at 2982
  Illegal SpeI site found at 3327
  Illegal PstI site found at 3341
  Illegal NotI site found at 3310
  Illegal NotI site found at 3334
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 3304
  Illegal BglII site found at 2873
  Illegal BglII site found at 2933
  Illegal XhoI site found at 1026
  Illegal XhoI site found at 2052
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal prefix found at 3304
  Illegal suffix found at 3327
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal prefix found at 3304
  Plasmid lacks a suffix.
  Illegal XbaI site found at 3319
  Illegal SpeI site found at 3327
  Illegal PstI site found at 3341
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.