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Plasmid Files

pYES3/CT

High-copy episomal vector with a TRP1 marker for galactose-inducible expression of C-terminally tagged proteins in S. cerevisiae.

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pYES3 CT Sequence and MappYES3 CT.dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Technologies)
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 NaeI (5725) NgoMIV (5723) DraIII (5622) AanI - AanI - PsiI (5494) SwaI (5397) SnaBI (5093) NsiI (4741) HpaI (4448) BmgBI (4388) PfoI * (4346) NruI (4042) BlpI (4006) StuI (3972) Bpu10I (3948) BtgI (3895) BspDI - ClaI (3862) AarI - BfuAI - BspMI (3565) Bsu36I (3374) MfeI (3272) BstZ17I (3099) PmlI (3092) TRP1 promoter AatII (2958) PvuII (471) T7 promoter HindIII (501) Acc65I (507) KpnI (511) Eco53kI (515) SacI (517) BamHI (519) BstXI (546) EcoRI (550) BstXI (572) NotI (577) PaeR7I - PspXI - XhoI (583) PspOMI (595) ApaI (599) PmeI (682) CYC1 terminator BsrGI (874) BspQI - SapI (1027) PciI (1143) AlwNI (1559) AhdI (2036) BglI (2156) ScaI (2516) ZraI (2956) pYES3/CT 5870 bp
NaeI  (5725)
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.
NgoMIV  (5723)
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.
DraIII  (5622)
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.
AanI  (5494)
1 site
T T A T A A A A T A T T
AanI  (5494)
1 site
T T A T A A A A T A T T
PsiI  (5494)
1 site
T T A T A A A A T A T T
SwaI  (5397)
1 site
A T T T A A A T T A A A T T T A

SwaI is typically used at 25°C, but is 50% active at 37°C.
SnaBI  (5093)
1 site
T A C G T A A T G C A T
NsiI  (4741)
1 site
A T G C A T T A C G T A
HpaI  (4448)
1 site
G T T A A C C A A T T G
BmgBI  (4388)
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  (4346)
1 site
T C C N G G A A G G N C C T
* Blocked by Dcm methylation.
Sticky ends from different PfoI sites may not be compatible.
NruI  (4042)
1 site
T C G C G A A G C G C T
BlpI  (4006)
1 site
G C T N A G C C G A N T C G

Sticky ends from different BlpI sites may not be compatible.
StuI  (3972)
1 site
A G G C C T T C C G G A
Bpu10I  (3948)
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.
BtgI  (3895)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
BspDI  (3862)
1 site
A T C G A T T A G C T A
ClaI  (3862)
1 site
A T C G A T T A G C T A
AarI  (3565)
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  (3565)
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  (3565)
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.
Bsu36I  (3374)
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.
MfeI  (3272)
1 site
C A A T T G G T T A A C
BstZ17I  (3099)
1 site
G T A T A C C A T A T G
PmlI  (3092)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
AatII  (2958)
1 site
G A C G T C C T G C A G
PvuII  (471)
1 site
C A G C T G G T C G A C
HindIII  (501)
1 site
A A G C T T T T C G A A
Acc65I  (507)
1 site
G G T A C C C C A T G G
KpnI  (511)
1 site
G G T A C C C C A T G G
Eco53kI  (515)
1 site
G A G C T C C T C G A G
SacI  (517)
1 site
G A G C T C C T C G A G
BamHI  (519)
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.
BstXI  (546)
2 sites
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.
EcoRI  (550)
1 site
G A A T T C C T T A A G
BstXI  (572)
2 sites
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.
NotI  (577)
1 site
G C G G C C G C C G C C G G C G
PaeR7I  (583)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (583)
1 site
V C T C G A G B B G A G C T C V
XhoI  (583)
1 site
C T C G A G G A G C T C
PspOMI  (595)
1 site
G G G C C C C C C G G G
ApaI  (599)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PmeI  (682)
1 site
G T T T A A A C C A A A T T T G
BsrGI  (874)
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.
BspQI  (1027)
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  (1027)
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.
PciI  (1143)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AlwNI  (1559)
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  (2036)
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.
BglI  (2156)
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.
ScaI  (2516)
1 site
A G T A C T T C A T G A
ZraI  (2956)
1 site
G A C G T C C T G C A G
2μ ori
4449 .. 5329  =  881 bp
yeast 2μ plasmid origin of replication
2μ ori
4449 .. 5329  =  881 bp
yeast 2μ plasmid origin of replication
AmpR
1963 .. 2823  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1963 .. 2754  =  792 bp
   263 amino acids  =  29.0 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1963 .. 2823  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   2755 .. 2823  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1963 .. 2823  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
TRP1
3133 .. 3807  =  675 bp
224 amino acids  =  24.1 kDa
Product: phosphoribosylanthranilate isomerase,
required for tryptophan biosynthesis
yeast auxotrophic marker
TRP1
3133 .. 3807  =  675 bp
224 amino acids  =  24.1 kDa
Product: phosphoribosylanthranilate isomerase,
required for tryptophan biosynthesis
yeast auxotrophic marker
ori
1204 .. 1792  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1204 .. 1792  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
5398 .. 5853  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
5398 .. 5853  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
GAL1 promoter
2 .. 443  =  442 bp
inducible promoter, regulated by Gal4
GAL1 promoter
2 .. 443  =  442 bp
inducible promoter, regulated by Gal4
CYC1 terminator
709 .. 956  =  248 bp
transcription terminator for CYC1
CYC1 terminator
709 .. 956  =  248 bp
transcription terminator for CYC1
AmpR promoter
2824 .. 2928  =  105 bp
AmpR promoter
2824 .. 2928  =  105 bp
TRP1 promoter
3031 .. 3132  =  102 bp
TRP1 promoter
3031 .. 3132  =  102 bp
MCS
501 .. 600  =  100 bp
multiple cloning site
MCS
501 .. 600  =  100 bp
multiple cloning site
V5 tag
607 .. 648  =  42 bp
14 amino acids  =  1.4 kDa
Product: epitope tag from simian virus 5
V5 tag
607 .. 648  =  42 bp
14 amino acids  =  1.4 kDa
Product: epitope tag from simian virus 5
T7 promoter
475 .. 493  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
475 .. 493  =  19 bp
promoter for bacteriophage T7 RNA polymerase
6xHis
658 .. 675  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
658 .. 675  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
UAS
2 .. 119  =  118 bp
upstream activating sequence mediating
Gal4-dependent induction
UAS
2 .. 119  =  118 bp
upstream activating sequence mediating
Gal4-dependent induction
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