Part:BBa_K3271007

His3::NatMX-EXSP pSB1K3

The prefix and suffix (EXSP) of the pSB1K3 backbone are flanked by the upstream and downstream homology sequences of the His3 locus from S. cerevisiae, respectively. Between the upstream homology and the prefix is the NatMX6 selection marker which confers resistance to nourseothricin in S. cerevisiae. To target a DNA part of interest to the His3 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 His3 locus upstream homology along with NatMX resistance 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 nourseothricin selection.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal prefix found at 3414
  Illegal suffix found at 3436
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 3414
  Illegal SpeI site found at 3437
  Illegal PstI site found at 3451
  Illegal NotI site found at 3420
  Illegal NotI site found at 3444
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 3414
  Illegal XhoI site found at 1026
  Illegal XhoI site found at 2052
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal prefix found at 3414
  Illegal suffix found at 3437
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal prefix found at 3414
  Plasmid lacks a suffix.
  Illegal XbaI site found at 3429
  Illegal SpeI site found at 3437
  Illegal PstI site found at 3451
  Illegal NgoMIV site found at 2945
  Illegal NgoMIV site found at 3034
  Illegal AgeI site found at 2663
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.