Difference between revisions of "Part:BBa K165062"

Revision as of 00:41, 30 October 2008

pRS305 yeast shuttle vector, LEU2 selection

Useful for chromosomal integration of a device into yeast strains. This will insert the vector at the specific locus of the LEU2 gene. To be used in conjunction with leucine drop-out media for positive selection of transformed cells. Transformation using this vector requires linearization of the plasmid by cutting with BstEII.

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

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

Vector: NotI, PstI Prefix: NotI,SpeI Suffix: XbaI, PstI

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 1116
Illegal EcoRI site found at 3178
Illegal XbaI site found at 3148
Illegal SpeI site found at 3154
Illegal PstI site found at 3172
12
INCOMPATIBLE WITH RFC[12]
Illegal EcoRI site found at 1116
Illegal EcoRI site found at 3178
Illegal SpeI site found at 3154
Illegal PstI site found at 3172
Illegal NotI site found at 3140
21
INCOMPATIBLE WITH RFC[21]
Illegal EcoRI site found at 1116
Illegal EcoRI site found at 3178
Illegal BamHI site found at 3160
Illegal XhoI site found at 3211
23
INCOMPATIBLE WITH RFC[23]
Illegal EcoRI site found at 1116
Illegal EcoRI site found at 3178
Illegal XbaI site found at 3148
Illegal SpeI site found at 3154
Illegal PstI site found at 3172
25
INCOMPATIBLE WITH RFC[25]
Illegal EcoRI site found at 1116
Illegal EcoRI site found at 3178
Illegal XbaI site found at 3148
Illegal SpeI site found at 3154
Illegal PstI site found at 3172
Illegal NgoMIV site found at 2799
Illegal AgeI site found at 1503
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 4584
Illegal SapI site found at 3501

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 __NOTOC__
 <partinfo>BBa_K165062 short</partinfo>
-Useful for chromosomal integration of a part into yeast strains.  Insertion of a single Biobrick (or biofusion) part requires cutting with XbaI and PstI.  Another option for this vector is to insert two Biobrick/biofusion parts by cutting the first part with NotI and SpeI and the second part with XbaI and PstI, while cutting the vector with NotI and PstI.  Note that this part does not conform to Standard Assembly requirements and will maintain idempotency.  Once a construct of interest has been inserted into the vector, it should be linearized with a digest using BstEII.  This vector will supplement a LEU2 auxotrophic yeast strain.  The part is then transformed into yeast according to the protocol outlined in this paper:
+Useful for chromosomal integration of a device into yeast strains.  This will insert the vector at the specific locus of the LEU2 gene.  To be used in conjunction with leucine drop-out media for positive selection of transformed cells.  Transformation using this vector requires linearization of the plasmid by cutting with BstEII.
+To incorporate a single Biobrick part into this vector for subsequent transformation into yeast, one should cut both vector and part with XbaI and PstI.
+Alternatively, one can do the final ligation step between two Biobrick parts into this vector by cutting with the following:
-R Daniel Gietz & Robert H Schiestl. High-efficiency yeast transformation using the LiAc/SS carrier DNA/PEG method . Nature protocols
+Vector: NotI, PstI
+Prefix: NotI,SpeI
+Suffix: XbaI, PstI
+Yeast transformation protocol:
+R Daniel Gietz & Robert H Schiestl. High-efficiency yeast transformation using the LiAc/SS carrier DNA/PEG method . Nature protocols (2007)
 <!-- Add more about the biology of this part here