pESC-TRP

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

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) BfuAI - BspMI - PaqCI (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
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).
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.
PaqCI  (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 PaqCI recognition sequence.
Sticky ends from different PaqCI sites may not be compatible.
Cleavage can be improved with PaqCI Activator.
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
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

Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
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
2 segments
   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
2 segments
   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
ORF:  718 .. 999  =  282 bp
ORF:  93 amino acids  =  10.8 kDa
ORF:  4253 .. 4519  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  5150 .. 5440  =  291 bp
ORF:  96 amino acids  =  11.5 kDa
ORF:  468 .. 1142  =  675 bp
ORF:  224 amino acids  =  24.1 kDa
ORF:  4123 .. 4983  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  1479 .. 1790  =  312 bp
ORF:  103 amino acids  =  11.3 kDa
ORF:  5916 .. 6236  =  321 bp
ORF:  106 amino acids  =  12.9 kDa
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Download pESC-TRP.dna file