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Plasmid Files

pESC-TRP

Yeast episomal vector with a TRP1 marker, for galactose-regulated expression and tagging of up to two genes.

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pESC-TRP Sequence and MappESC-TRP.dna
Map and Sequence File   
Sequence Author:  Agilent Technologies
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 XcmI (6405) BmgBI (6194) NsiI (5404) ScaI (4676) BpmI (4266) BsaI (4257) AhdI (4196) AlwNI (3719) PciI (3303) PfoI (46) BspEI * (332) PmlI (427) Bsu36I (709) EcoRV (752) BsgI (880) AarI - BfuAI - BspMI (900) DraIII (1465) NgoMIV (1566) NaeI (1568) PacI (2078) Eco53kI (2084) SacI (2086) BglII (2088) BspDI - ClaI (2118) SpeI (2123) EagI - NotI (2130) EcoRI (2152) BstBI (2156) AgeI (2447) BamHI (2830) PspOMI (2854) TspMI - XmaI (2857) ApaI (2858) SmaI - SrfI (2859) SalI (2864) PaeR7I - PspXI - XhoI (2900) Acc65I (2913) BtgI - KpnI (2917) SacII (2920) NheI (2922) BmtI (2926) PpuMI (2977) BsrGI (3051) MluI (3058) BspQI - SapI (3187) pESC-TRP 6525 bp
XcmI  (6405)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
BmgBI  (6194)
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.
NsiI  (5404)
1 site
A T G C A T T A C G T A
ScaI  (4676)
1 site
A G T A C T T C A T G A
BpmI  (4266)
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.
BsaI  (4257)
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  (4196)
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  (3719)
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.
PciI  (3303)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PfoI  (46)
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.
BspEI  (332)
1 site
T C C G G A A G G C C T
* Blocked by Dam methylation.
PmlI  (427)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
Bsu36I  (709)
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.
EcoRV  (752)
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.
BsgI  (880)
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).
AarI  (900)
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.
BfuAI  (900)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BfuAI
recognition sequence.
Sticky ends from different BfuAI sites may not be compatible.
BfuAI is typically used at 50°C, but is 50% active at 37°C.
BspMI  (900)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BspMI
recognition sequence.
Sticky ends from different BspMI sites may not be compatible.
DraIII  (1465)
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.
NgoMIV  (1566)
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  (1568)
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.
PacI  (2078)
1 site
T T A A T T A A A A T T A A T T
Eco53kI  (2084)
1 site
G A G C T C C T C G A G
SacI  (2086)
1 site
G A G C T C C T C G A G
BglII  (2088)
1 site
A G A T C T T C T A G A
BspDI  (2118)
1 site
A T C G A T T A G C T A
ClaI  (2118)
1 site
A T C G A T T A G C T A
SpeI  (2123)
1 site
A C T A G T T G A T C A
EagI  (2130)
1 site
C G G C C G G C C G G C
NotI  (2130)
1 site
G C G G C C G C C G C C G G C G
EcoRI  (2152)
1 site
G A A T T C C T T A A G
BstBI  (2156)
1 site
T T C G A A A A G C T T
AgeI  (2447)
1 site
A C C G G T T G G C C A

AgeI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
BamHI  (2830)
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.
PspOMI  (2854)
1 site
G G G C C C C C C G G G
TspMI  (2857)
1 site
C C C G G G G G G C C C
XmaI  (2857)
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.
ApaI  (2858)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
SmaI  (2859)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
SrfI  (2859)
1 site
G C C C G G G C C G G G C C C G
SalI  (2864)
1 site
G T C G A C C A G C T G
PaeR7I  (2900)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (2900)
1 site
V C T C G A G B B G A G C T C V
XhoI  (2900)
1 site
C T C G A G G A G C T C
Acc65I  (2913)
1 site
G G T A C C C C A T G G
BtgI  (2917)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
KpnI  (2917)
1 site
G G T A C C C C A T G G
SacII  (2920)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII
recognition sequence.
NheI  (2922)
1 site
G C T A G C C G A T C G
BmtI  (2926)
1 site
G C T A G C C G A T C G
PpuMI  (2977)
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.
BsrGI  (3051)
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.
MluI  (3058)
1 site
A C G C G T T G C G C A
BspQI  (3187)
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  (3187)
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
5115 .. 6457  =  1343 bp
yeast 2μ plasmid origin of replication
2μ ori
5115 .. 6457  =  1343 bp
yeast 2μ plasmid origin of replication
AmpR
4123 .. 4983  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   4123 .. 4914  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4123 .. 4983  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   4915 .. 4983  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4123 .. 4983  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
TRP1
468 .. 1142  =  675 bp
224 amino acids  =  24.1 kDa
Product: phosphoribosylanthranilate isomerase,
required for tryptophan biosynthesis
yeast auxotrophic marker
TRP1
468 .. 1142  =  675 bp
224 amino acids  =  24.1 kDa
Product: phosphoribosylanthranilate isomerase,
required for tryptophan biosynthesis
yeast auxotrophic marker
GAL1,10 promoter
2162 .. 2826  =  665 bp
divergent inducible promoter, regulated by Gal4
GAL1,10 promoter
2162 .. 2826  =  665 bp
divergent inducible promoter, regulated by Gal4
ori
3364 .. 3952  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3364 .. 3952  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
1241 .. 1696  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
1241 .. 1696  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
TRP1 promoter
187 .. 467  =  281 bp
TRP1 promoter
187 .. 467  =  281 bp
CYC1 terminator
2932 .. 3121  =  190 bp
transcription terminator for CYC1
CYC1 terminator
2932 .. 3121  =  190 bp
transcription terminator for CYC1
ADH1 terminator
1779 .. 1944  =  166 bp
transcription terminator for the S. cerevisiae alcohol
dehydrogenase 1 (ADH1) gene
ADH1 terminator
1779 .. 1944  =  166 bp
transcription terminator for the S. cerevisiae alcohol
dehydrogenase 1 (ADH1) gene
AmpR promoter
4984 .. 5088  =  105 bp
AmpR promoter
4984 .. 5088  =  105 bp
MCS 2
2830 .. 2927  =  98 bp
multiple cloning site 2
MCS 2
2830 .. 2927  =  98 bp
multiple cloning site 2
MCS 1
2074 .. 2160  =  87 bp
multiple cloning site 1
MCS 1
2074 .. 2160  =  87 bp
multiple cloning site 1
UAS
2378 .. 2495  =  118 bp
upstream activating sequence mediating
Gal4-dependent induction
UAS
2378 .. 2495  =  118 bp
upstream activating sequence mediating
Gal4-dependent induction
Myc
2873 .. 2902  =  30 bp
10 amino acids  =  1.2 kDa
Product: Myc (human c-Myc oncogene) epitope tag
Myc
2873 .. 2902  =  30 bp
10 amino acids  =  1.2 kDa
Product: Myc (human c-Myc oncogene) epitope tag
FLAG
2092 .. 2115  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG® epitope tag, followed by an
enterokinase cleavage site
FLAG
2092 .. 2115  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG® epitope tag, followed by an
enterokinase cleavage site
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