Plasmid

Part:BBa_K165063

Designed by: Aaron Glieberman   Group: iGEM08_BrownTwo   (2008-10-29)
Revision as of 00:48, 30 October 2008 by Aaron Glieberman (Talk | contribs)

pRS306 yeast shuttle vector, URA3 selection

Ampicillin resistance when grown in bacteria. This plasmid is not currently Biobrick compatible, but is useful for inserting a final construct into a yeast cell.

Useful for chromosomal integration of a device into yeast strains. This will insert the vector at the specific locus of the URA3 gene. To be used in conjunction with uracil 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. Sikorski and P. Hieter, Genetics, 122: 19-27 (1989).


Yeast transformation protocol: R Daniel Gietz & Robert H Schiestl. 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 2047
    Illegal XbaI site found at 2017
    Illegal SpeI site found at 2023
    Illegal PstI site found at 389
    Illegal PstI site found at 2041
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 2047
    Illegal SpeI site found at 2023
    Illegal PstI site found at 389
    Illegal PstI site found at 2041
    Illegal NotI site found at 2009
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 2047
    Illegal BamHI site found at 2029
    Illegal XhoI site found at 2080
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 2047
    Illegal XbaI site found at 2017
    Illegal SpeI site found at 2023
    Illegal PstI site found at 389
    Illegal PstI site found at 2041
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 2047
    Illegal XbaI site found at 2017
    Illegal SpeI site found at 2023
    Illegal PstI site found at 389
    Illegal PstI site found at 2041
    Illegal NgoMIV site found at 1668
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1079
    Illegal BsaI.rc site found at 3453
    Illegal SapI site found at 2370
    Illegal SapI.rc site found at 926


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