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Plasmid Files

pAD-GAL4

Phagemid vector carrying the GAL4 activation domain for yeast two-hybrid analysis.

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pAD-GAL4 Sequence and MappAD-GAL4.dna
Map and Sequence File   
Sequence Author:  I.M.A.G.E. Consortium
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 SnaBI (6978) BmgBI (6536) PfoI (6315) AatII (6204) ZraI (6202) ScaI (5762) PvuI (5652) BsaI (5343) AhdI (5282) AlwNI (4805) BspQI - SapI (4273) EagI - NotI (4201) DraIII (3962) BtgZI (3954) NaeI (3856) BsgI (352) ATG Acc65I (479) KpnI (483) RsrII (557) MluI (726) EcoRI (833) SrfI (842) PaeR7I - PspXI - XhoI (847) SalI (858) XbaI (864) T7 promoter PstI (895) BglII (897) MscI (1033) BsrGI (1551) EcoRV (1926) AflII (2357) BspDI - ClaI (2522) BstEII (2637) PflMI (2766) PpuMI (2782) SfiI (3303) NcoI (3342) Bsu36I (3592) NgoMIV (3854) pAD-GAL4 7620 bp
SnaBI  (6978)
1 site
T A C G T A A T G C A T
BmgBI  (6536)
1 site
C A C G T C G T G C A G

This recognition sequence is asymmetric, so ligating blunt ends
generated by BmgBI will not always regenerate a BmgBI site.
PfoI  (6315)
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.
AatII  (6204)
1 site
G A C G T C C T G C A G
ZraI  (6202)
1 site
G A C G T C C T G C A G
ScaI  (5762)
1 site
A G T A C T T C A T G A
PvuI  (5652)
1 site
C G A T C G G C T A G C
BsaI  (5343)
1 site
G G T C T C N C C A G A G N ( N ) 4

Sticky ends from different BsaI sites may not be compatible.
BsaI can be used between 37°C and 50°C.
AhdI  (5282)
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.
AlwNI  (4805)
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.
BspQI  (4273)
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  (4273)
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.
EagI  (4201)
1 site
C G G C C G G C C G G C
NotI  (4201)
1 site
G C G G C C G C C G C C G G C G
DraIII  (3962)
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.
BtgZI  (3954)
1 site
G C G A T G ( N ) 10 C G C T A C ( N ) 10 ( N ) 4

Sticky ends from different BtgZI sites may not be compatible.
After cleavage, BtgZI can remain bound to DNA and alter its
electrophoretic mobility.
BtgZI is typically used at 60°C, but is 75% active at 37°C.
NaeI  (3856)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI
recognition sequence.
BsgI  (352)
1 site
G T G C A G ( N ) 14 N N C A C G T C ( N ) 14

Efficient cleavage requires at least two copies of the BsgI
recognition sequence.
Sticky ends from different BsgI sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
Acc65I  (479)
1 site
G G T A C C C C A T G G
KpnI  (483)
1 site
G G T A C C C C A T G G
RsrII  (557)
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.
MluI  (726)
1 site
A C G C G T T G C G C A
EcoRI  (833)
1 site
G A A T T C C T T A A G
SrfI  (842)
1 site
G C C C G G G C C G G G C C C G
PaeR7I  (847)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (847)
1 site
V C T C G A G B B G A G C T C V
XhoI  (847)
1 site
C T C G A G G A G C T C
SalI  (858)
1 site
G T C G A C C A G C T G
XbaI  (864)
1 site
T C T A G A A G A T C T
PstI  (895)
1 site
C T G C A G G A C G T C
BglII  (897)
1 site
A G A T C T T C T A G A
MscI  (1033)
1 site
T G G C C A A C C G G T
BsrGI  (1551)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
EcoRV  (1926)
1 site
G A T A T C C T A T A G

EcoRV is reportedly more prone than its isoschizomer Eco32I to
delete a base after cleavage.
AflII  (2357)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
BspDI  (2522)
1 site
A T C G A T T A G C T A
ClaI  (2522)
1 site
A T C G A T T A G C T A
BstEII  (2637)
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  (2766)
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  (2782)
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.
SfiI  (3303)
1 site
G G C C N N N N N G G C C C C G G N N N N N C C G G

Efficient cleavage requires at least two copies of the SfiI
recognition sequence.
Sticky ends from different SfiI sites may not be compatible.
NcoI  (3342)
1 site
C C A T G G G G T A C C
Bsu36I  (3592)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
NgoMIV  (3854)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV
recognition sequence.
2μ ori
6456 .. 7620  =  1165 bp
yeast 2μ plasmid origin of replication
2μ ori
6456 .. 7620  =  1165 bp
yeast 2μ plasmid origin of replication
LEU2
1582 .. 2676  =  1095 bp
364 amino acids  =  39.0 kDa
Product: 3-isopropylmalate dehydrogenase, required
for leucine biosynthesis
yeast auxotrophic marker
LEU2
1582 .. 2676  =  1095 bp
364 amino acids  =  39.0 kDa
Product: 3-isopropylmalate dehydrogenase, required
for leucine biosynthesis
yeast auxotrophic marker
AmpR
5209 .. 6069  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   5209 .. 6000  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
5209 .. 6069  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   6001 .. 6069  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
5209 .. 6069  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
4450 .. 5038  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
4450 .. 5038  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
3729 .. 4184  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
3729 .. 4184  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
LEU2 promoter
2677 .. 3081  =  405 bp
LEU2 promoter
2677 .. 3081  =  405 bp
ADH1 promoter
5 .. 406  =  402 bp
promoter for alcohol dehydrogenase 1
ADH1 promoter
5 .. 406  =  402 bp
promoter for alcohol dehydrogenase 1
GAL4 activation domain
488 .. 829  =  342 bp
114 amino acids  =  12.4 kDa
Product: activation domain of the GAL4
transcriptional activator
GAL4 activation domain
488 .. 829  =  342 bp
114 amino acids  =  12.4 kDa
Product: activation domain of the GAL4
transcriptional activator
ADH1 terminator
1277 .. 1464  =  188 bp
transcription terminator for alcohol dehydrogenase 1
ADH1 terminator
1277 .. 1464  =  188 bp
transcription terminator for alcohol dehydrogenase 1
AmpR promoter
6070 .. 6174  =  105 bp
AmpR promoter
6070 .. 6174  =  105 bp
MCS
833 .. 869  =  37 bp
multiple cloning site
MCS
833 .. 869  =  37 bp
multiple cloning site
SV40 NLS
452 .. 472  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
SV40 NLS
452 .. 472  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
T7 promoter
872 .. 890  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
872 .. 890  =  19 bp
promoter for bacteriophage T7 RNA polymerase
ATG
422 .. 424  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
422 .. 424  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
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