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Plasmid Files

pOAD

Yeast two-hybrid "prey" vector for fusing a gene to the GAL4 activation domain.

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pOAD Sequence and MappOAD.dna
Map and Sequence File   
Sequence Author:  Fields Lab
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 NcoI (8423) PvuII (8415) EcoRI (8407) MluI (8300) SexAI * (8271) AfeI (8168) RsrII (8131) KpnI (8057) Acc65I (8053) PacI (7195) BsmI (6835) Bpu10I (6801) PfoI (6531) BspQI - SapI (6465) DrdI (6246) AlwNI (5939) AhdI (5460) AatII (4541) ZraI (4539) TspMI - XmaI (5) SmaI (7) SalI (16) PstI (26) MscI (164) SpeI (912) PflFI - Tth111I (1304) DraIII (1790) NheI (2163) BmtI (2167) BanII (2598) AarI (2665) AflII (3726) BstEII (4006) PflMI (4147) PpuMI (4163) pOAD 8429 bp
NcoI  (8423)
1 site
C C A T G G G G T A C C
PvuII  (8415)
1 site
C A G C T G G T C G A C
EcoRI  (8407)
1 site
G A A T T C C T T A A G
MluI  (8300)
1 site
A C G C G T T G C G C A
SexAI  (8271)
1 site
A C C W G G T T G G W C C A
* Blocked by Dcm methylation.
Sticky ends from different SexAI sites may not be compatible.
AfeI  (8168)
1 site
A G C G C T T C G C G A
RsrII  (8131)
1 site
C G G W C C G G C C W G G C

Efficient cleavage requires at least two copies of the RsrII
recognition sequence.
Sticky ends from different RsrII sites may not be compatible.
For full activity, add fresh DTT.
KpnI  (8057)
1 site
G G T A C C C C A T G G
Acc65I  (8053)
1 site
G G T A C C C C A T G G
PacI  (7195)
1 site
T T A A T T A A A A T T A A T T
BsmI  (6835)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
Bpu10I  (6801)
1 site
C C T N A G C G G A N T C G

Efficient cleavage requires at least two copies of the Bpu10I
recognition sequence.
This recognition sequence is asymmetric, so ligating sticky ends
generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
PfoI  (6531)
1 site
T C C N G G A A G G N C C T

Sticky ends from different PfoI sites may not be compatible.
BspQI  (6465)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (6465)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be
mixed before removing an aliquot.
DrdI  (6246)
1 site
G A C N N N N N N G T C C T G N N N N N N C A G

Sticky ends from different DrdI sites may not be compatible.
AlwNI  (5939)
1 site
C A G N N N C T G G T C N N N G A C

Sticky ends from different AlwNI sites may not be compatible.
AhdI  (5460)
1 site
G A C N N N N N G T C C T G N N N N N C A G

The 1-base overhangs produced by AhdI may be hard to ligate.
Sticky ends from different AhdI sites may not be compatible.
AatII  (4541)
1 site
G A C G T C C T G C A G
ZraI  (4539)
1 site
G A C G T C C T G C A G
TspMI  (5)
1 site
C C C G G G G G G C C C
XmaI  (5)
1 site
C C C G G G G G G C C C

Efficient cleavage requires at least two copies of the XmaI
recognition sequence.
Full cleavage with XmaI may require a long incubation.
SmaI  (7)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
SalI  (16)
1 site
G T C G A C C A G C T G
PstI  (26)
1 site
C T G C A G G A C G T C
MscI  (164)
1 site
T G G C C A A C C G G T
SpeI  (912)
1 site
A C T A G T T G A T C A
PflFI  (1304)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (1304)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
DraIII  (1790)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
NheI  (2163)
1 site
G C T A G C C G A T C G
BmtI  (2167)
1 site
G C T A G C C G A T C G
BanII  (2598)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
AarI  (2665)
1 site
C A C C T G C ( N ) 4 G T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the AarI
recognition sequence.
Sticky ends from different AarI sites may not be compatible.
After cleavage, AarI can remain bound to DNA and alter its
electrophoretic mobility.
AflII  (3726)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
BstEII  (4006)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
PflMI  (4147)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
PpuMI  (4163)
1 site
R G G W C C Y Y C C W G G R

Sticky ends from different PpuMI sites may not be compatible.
CEN/ARS
1427 .. 2589  =  1163 bp
S. cerevisiae CEN4 centromere fused to the
autonomously replicating sequence ARS1/ARS416
CEN/ARS
1427 .. 2589  =  1163 bp
S. cerevisiae CEN4 centromere fused to the
autonomously replicating sequence ARS1/ARS416
LEU2
2951 .. 4045  =  1095 bp
364 amino acids  =  39.0 kDa
Product: 3-isopropylmalate dehydrogenase, required
for leucine biosynthesis
yeast auxotrophic marker
LEU2
2951 .. 4045  =  1095 bp
364 amino acids  =  39.0 kDa
Product: 3-isopropylmalate dehydrogenase, required
for leucine biosynthesis
yeast auxotrophic marker
AmpR
4673 .. 5533  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   4673 .. 4741  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4673 .. 5533  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   4742 .. 5533  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4673 .. 5533  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ADH1 promoter
7281 .. 7981  =  701 bp
promoter for alcohol dehydrogenase 1
ADH1 promoter
7281 .. 7981  =  701 bp
promoter for alcohol dehydrogenase 1
ori
5704 .. 6292  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin
of replication
ori
5704 .. 6292  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin
of replication
LEU2 promoter
4058 .. 4462  =  405 bp
LEU2 promoter
4058 .. 4462  =  405 bp
GAL4 activation domain
8062 .. 8403  =  342 bp
114 amino acids  =  12.4 kDa
Product: activation domain of the GAL4
transcriptional activator
GAL4 activation domain
8062 .. 8403  =  342 bp
114 amino acids  =  12.4 kDa
Product: activation domain of the GAL4
transcriptional activator
ADH1 terminator
408 .. 595  =  188 bp
transcription terminator for alcohol dehydrogenase 1
ADH1 terminator
408 .. 595  =  188 bp
transcription terminator for alcohol dehydrogenase 1
AmpR promoter
4568 .. 4672  =  105 bp
AmpR promoter
4568 .. 4672  =  105 bp
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