Part:BBa_K3271005

Ade4::Ura3-EXSP pSB1K3

The prefix and suffix (EXSP) of the pSB1K3 backbone are flanked by the upstream and downstream homology sequences of the Ade4 locus from S. cerevisiae, respectively. Between the upstream homology and the prefix is the Ura3 auxotrophic selection marker. To target a DNA part of interest to the Ade4 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 Ura3 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 Ura-.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal prefix found at 3530
  Illegal suffix found at 3552
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 3530
  Illegal SpeI site found at 3553
  Illegal PstI site found at 3567
  Illegal NotI site found at 3536
  Illegal NotI site found at 3560
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 3530
  Illegal XhoI site found at 1026
  Illegal XhoI site found at 2052
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal prefix found at 3530
  Illegal suffix found at 3553
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal prefix found at 3530
  Plasmid lacks a suffix.
  Illegal XbaI site found at 3545
  Illegal SpeI site found at 3553
  Illegal PstI site found at 3567
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal BsaI site found at 3212
  Illegal SapI.rc site found at 3059