pBridge

Yeast three-hybrid vector for expressing a "bait" protein fused to the GAL4 DNA-binding domain, plus an additional bridge protein.

Sequence Author: Clontech (TaKaRa)

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NsiI (6241) PfoI (5222) AatII (5111) ZraI (5109) ScaI (4669) TsoI (4588) PvuI (4559) BglI (4309) BpmI (4259) AhdI (4189) AlwNI (3712) AflIII - PciI (3296) PaeR7I - XhoI (651) HpaI (711) BsrGI (724) EcoRI (878) TspMI - XmaI (883) SmaI (885) BamHI (888) SalI (894) PstI (904) PsiI (963) NgoMIV (1112) NaeI (1114) BanII (1331) PaqCI (1398) BstXI (1541) Bsu36I (1590) XbaI (1747) BstAPI (1759) BstBI (2240) Bpu10I (2248) KasI (2373) NarI (2374) SfoI (2375) PluTI (2377) PflMI (2609) NdeI (2617) ATG PpuMI (2667) SV40 NLS EagI - NotI (2701) BglII (2718) BspDI - ClaI (2842) BseRI (2987) BspQI - SapI (3180) pBridge™ 6527 bp
NsiI  (6241)
1 site
A T G C A T T A C G T A
PfoI  (5222)
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  (5111)
1 site
G A C G T C C T G C A G
ZraI  (5109)
1 site
G A C G T C C T G C A G
ScaI  (4669)
1 site
A G T A C T T C A T G A
TsoI  (4588)
1 site
T A R C C A ( N ) 9 N N A T Y G G T ( N ) 9

Sticky ends from different TsoI sites may not be compatible.
After cleavage, TsoI can remain bound to DNA and alter its electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
PvuI  (4559)
1 site
C G A T C G G C T A G C
BglI  (4309)
1 site
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
BpmI  (4259)
1 site
C T G G A G ( N ) 14 N N G A C C T C ( N ) 14

Efficient cleavage requires at least two copies of the BpmI recognition sequence.
Sticky ends from different BpmI sites may not be compatible.
After cleavage, BpmI can remain bound to DNA and alter its electrophoretic mobility.
BpmI quickly loses activity at 37°C.
AhdI  (4189)
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  (3712)
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.
AflIII  (3296)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
PciI  (3296)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PaeR7I  (651)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (651)
1 site
C T C G A G G A G C T C
HpaI  (711)
1 site
G T T A A C C A A T T G
BsrGI  (724)
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.
EcoRI  (878)
1 site
G A A T T C C T T A A G
TspMI  (883)
1 site
C C C G G G G G G C C C
XmaI  (883)
1 site
C C C G G G G G G C C C

Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
SmaI  (885)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
BamHI  (888)
1 site
G G A T C C C C T A G G

After cleavage, BamHI-HF® (but not the original BamHI) can remain bound to DNA and alter its electrophoretic mobility.
SalI  (894)
1 site
G T C G A C C A G C T G
PstI  (904)
1 site
C T G C A G G A C G T C
PsiI  (963)
1 site
T T A T A A A A T A T T
NgoMIV  (1112)
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.
NaeI  (1114)
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.
BanII  (1331)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
PaqCI  (1398)
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 PaqCI recognition sequence.
Sticky ends from different PaqCI sites may not be compatible.
Cleavage can be improved with PaqCI Activator.
BstXI  (1541)
1 site
C C A N N N N N N T G G G G T N N N N N N A C C

Sticky ends from different BstXI sites may not be compatible.
Bsu36I  (1590)
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.
XbaI  (1747)
1 site
T C T A G A A G A T C T
BstAPI  (1759)
1 site
G C A N N N N N T G C C G T N N N N N A C G

Sticky ends from different BstAPI sites may not be compatible.
BstBI  (2240)
1 site
T T C G A A A A G C T T
Bpu10I  (2248)
1 site
C C T N A G C G G A N T C G

Cleavage may be enhanced when more than one copy of the Bpu10I recognition sequence is present.
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.
KasI  (2373)
1 site
G G C G C C C C G C G G
NarI  (2374)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the NarI recognition sequence.
SfoI  (2375)
1 site
G G C G C C C C G C G G
PluTI  (2377)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI recognition sequence.
PflMI  (2609)
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.
NdeI  (2617)
1 site
C A T A T G G T A T A C

Prolonged incubation with NdeI may lead to removal of additional nucleotides.
PpuMI  (2667)
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.
EagI  (2701)
1 site
C G G C C G G C C G G C
NotI  (2701)
1 site
G C G G C C G C C G C C G G C G
BglII  (2718)
1 site
A G A T C T T C T A G A
BspDI  (2842)
1 site
A T C G A T T A G C T A
ClaI  (2842)
1 site
A T C G A T T A G C T A
BseRI  (2987)
1 site
G A G G A G ( N ) 8 N N C T C C T C ( N ) 8

Sticky ends from different BseRI sites may not be compatible.
BseRI quickly loses activity at 37°C.
Prolonged incubation with BseRI may lead to degradation of the DNA.
BspQI  (3180)
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  (3180)
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.
2μ ori
5363 .. 6527  =  1165 bp
yeast 2μ plasmid origin of replication
2μ ori
5363 .. 6527  =  1165 bp
yeast 2μ plasmid origin of replication
AmpR
4116 .. 4976  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   4116 .. 4907  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4116 .. 4976  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   4908 .. 4976  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4116 .. 4976  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
TRP1
1161 .. 1835  =  675 bp
224 amino acids  =  24.1 kDa
Product: phosphoribosylanthranilate isomerase, required for tryptophan biosynthesis
yeast auxotrophic marker
TRP1
1161 .. 1835  =  675 bp
224 amino acids  =  24.1 kDa
Product: phosphoribosylanthranilate isomerase, re