YEp352

Yeast/E. coli shuttle vector with a URA3 marker. See also YEp351.
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lac operator AhdI (3952) BsrFI (3867) NmeAIII (3805) TsoI (3554) AatII (3033) ZraI (3031) EcoRI (15) Eco53kI (23) SacI (25) Acc65I (27) KpnI - TspMI - XmaI (31) SmaI (33) BamHI (36) XbaI (42) SalI (48) PstI - SbfI (58) BfuAI - BspMI (61) SphI (64) HindIII (66) KasI (230) NarI (231) SfoI (232) PluTI (234) PsiI (351) Bpu10I (433) StuI (730) PspOMI * (788) ApaI * (792) BsmI (806) BstBI (899) NcoI (957) EcoRV (978) PpuMI (1010) XcmI (1039) BsgI (1217) NdeI (1246) BstAPI (1533) HpaI (1613) BsaAI - SnaBI (2255) YEp352 5181 bp
AhdI  (3952)
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.
BsrFI  (3867)
1 site
R C C G G Y Y G G C C R

Cleavage may be enhanced when more than one copy of the BsrFI recognition sequence is present.
After cleavage, BsrFI can remain bound to DNA and alter its electrophoretic mobility.
NmeAIII  (3805)
1 site
G C C G A G ( N ) 18-19 N N C G G C T C ( N ) 18-19

Efficient cleavage requires at least two copies of the NmeAIII recognition sequence.
Sticky ends from different NmeAIII sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
TsoI  (3554)
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).
AatII  (3033)
1 site
G A C G T C C T G C A G
ZraI  (3031)
1 site
G A C G T C C T G C A G
EcoRI  (15)
1 site
G A A T T C C T T A A G
Eco53kI  (23)
1 site
G A G C T C C T C G A G
SacI  (25)
1 site
G A G C T C C T C G A G
Acc65I  (27)
1 site
G G T A C C C C A T G G
KpnI  (31)
1 site
G G T A C C C C A T G G
TspMI  (31)
1 site
C C C G G G G G G C C C
XmaI  (31)
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  (33)
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  (36)
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.
XbaI  (42)
1 site
T C T A G A A G A T C T
SalI  (48)
1 site
G T C G A C C A G C T G
PstI  (58)
1 site
C T G C A G G A C G T C
SbfI  (58)
1 site
C C T G C A G G G G A C G T C C
BfuAI  (61)
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  (61)
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.
SphI  (64)
1 site
G C A T G C C G T A C G
HindIII  (66)
1 site
A A G C T T T T C G A A
KasI  (230)
1 site
G G C G C C C C G C G G
NarI  (231)
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  (232)
1 site
G G C G C C C C G C G G
PluTI  (234)
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.
PsiI  (351)
1 site
T T A T A A A A T A T T
Bpu10I  (433)
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.
StuI  (730)
1 site
A G G C C T T C C G G A
PspOMI  (788)
1 site
G G G C C C C C C G G G
* Blocked by Dcm methylation.
ApaI  (792)
1 site
G G G C C C C C C G G G
* Blocked by Dcm methylation.
ApaI can be used between 25°C and 37°C.
BsmI  (806)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
BstBI  (899)
1 site
T T C G A A A A G C T T
NcoI  (957)
1 site
C C A T G G G G T A C C
EcoRV  (978)
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.
PpuMI  (1010)
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.
XcmI  (1039)
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.
BsgI  (1217)
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).
NdeI  (1246)
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.
BstAPI  (1533)
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.
HpaI  (1613)
1 site
G T T A A C C A A T T G
BsaAI  (2255)
1 site
Y A C G T R R T G C A Y
SnaBI  (2255)
1 site
T A C G T A A T G C A T
2μ ori
1614 .. 2778  =  1165 bp
yeast 2μ plasmid origin of replication
2μ ori
1614 .. 2778  =  1165 bp
yeast 2μ plasmid origin of replication
AmpR
3165 .. 4025  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   3165 .. 3233  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3165 .. 4025  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   3234 .. 4025  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3165 .. 4025  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
URA3
364 .. 1167  =  804 bp
267 amino acids  =  29.3 kDa
URA3
364 .. 1167  =  804 bp
267 amino acids  =  29.3 kDa
ori
4196 .. 4784  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
ori
4196 .. 4784  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
lacZα
1 .. 291  =  291 bp
96 amino acids  =  10.9 kDa
Product: LacZα fragment of β-galactosidase
lacZα
1 .. 291  =  291 bp
96 amino acids  =  10.9 kDa
Product: LacZα fragment of β-galactosidase
URA3 promoter
1168 .. 1378  =  211 bp
URA3 promoter
1168 .. 1378  =  211 bp
AmpR promoter
3060 .. 3164  =  105 bp
AmpR promoter
3060 .. 3164  =  105 bp
lac promoter
5108 .. 5138  =  31 bp
3 segments
   Segment 1:  -35  
   5108 .. 5113  =  6 bp
promoter for the E. coli lac operon
lac promoter
5108 .. 5138  =  31 bp
3 segments
   Segment 2:  
   5114 .. 5131  =  18 bp
promoter for the E. coli lac operon
lac promoter
5108 .. 5138  =  31 bp
3 segments
   Segment 3:  -10  
   5132 .. 5138  =  7 bp
promoter for the E. coli lac operon
lac promoter
5108 .. 5138  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
5146 .. 5162  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lac operator
5146 .. 5162  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
MCS
15 .. 71  =  57 bp
pUC19 multiple cloning site
MCS
15 .. 71  =  57 bp
pUC19 multiple cloning site
M13 fwd
75 .. 91  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
75 .. 91  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
5170 .. 5  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
5170 .. 5  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  1 .. 291  =  291 bp
ORF:  96 amino acids  =  10.9 kDa
ORF:  656 .. 1132  =  477 bp
ORF:  158 amino acids  =  16.8 kDa
ORF:  1649 .. 1939  =  291 bp
ORF:  96 amino acids  =  11.5 kDa
ORF:  3165 .. 4025  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  364 .. 1167  =  804 bp
ORF:  267 amino acids  =  29.3 kDa
ORF:  5109 .. 284  =  357 bp
ORF:  118 amino acids  =  13.4 kDa
ORF:  1395 .. 1634  =  240 bp
ORF:  79 amino acids  =  9.8 kDa
ORF:  2415 .. 2681  =  267 bp
ORF:  88 amino acids  =  10.3 kDa
ORF:  3629 .. 3895  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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