Part:BBa_K165061

pRS304* yeast shuttle vector, TRP1 selection

Ampicillin resistance when grown in bacteria. This plasmid is not compatible with Biobrick construction, but can be used for shuttling Biobrick parts into the yeast genome.

This integrating vector inserts at the specific locus of the TRP13 gene. To be used in conjunction with tryptophan drop-out media for positive selection of transformed cells. Transformation using this vector requires linearization of the plasmid by cutting with PstI.

To incorporate a single Biobrick part into this vector for subsequent transformation into yeast, one should cut both vector and part with EcoRI and SpeI.

Alternatively, one can do the final ligation step between two Biobrick parts into this vector by cutting with the following:

Vector: EcoRI, Not I

Prefix: EcoRI, SpeI

Suffix: XbaI, NotI

Original Sikorski vector was described in R. S. [http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pubmed&pubmedid=2659436 Sikorski and P. Hieter, Genetics, 122: 19-27 (1989)].

Vector NTI annotated sequence: File:PRS304star.gb

Yeast transformation protocol: R Daniel Gietz & Robert H Schiestl. [http://www.natureprotocols.com/2007/01/31/highefficiency_yeast_transform.php High-efficiency yeast transformation using the LiAc/SS carrier DNA/PEG method]. Nature protocols (2007)

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 1942
  Illegal XbaI site found at 550
  Illegal XbaI site found at 1912
  Illegal SpeI site found at 1918
  Illegal PstI site found at 251
  Illegal PstI site found at 1936
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 1942
  Illegal SpeI site found at 1918
  Illegal PstI site found at 251
  Illegal PstI site found at 1936
  Illegal NotI site found at 1904
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 1942
  Illegal BamHI site found at 1924
  Illegal XhoI site found at 1975
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 1942
  Illegal XbaI site found at 550
  Illegal XbaI site found at 1912
  Illegal SpeI site found at 1918
  Illegal PstI site found at 251
  Illegal PstI site found at 1936
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 1942
  Illegal XbaI site found at 550
  Illegal XbaI site found at 1912
  Illegal SpeI site found at 1918
  Illegal PstI site found at 251
  Illegal PstI site found at 1936
  Illegal NgoMIV site found at 1563
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 3348
  Illegal SapI site found at 2265