YIp351

Yeast/E. coli integrating shuttle vector with a LEU2 marker. See also YIp352.
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Eco53kI (23) lac operator BspQI - SapI (4182) DrdI (3963) PspFI (3765) BseYI (3761) AlwNI (3656) AhdI (3177) BsaI (3111) BpmI (3108) NmeAIII (3030) FspI (2954) ScaI (2696) BanII - SacI (25) AvaI - BsoBI - TspMI - XmaI (31) SmaI (33) BamHI (36) XbaI (42) SalI (48) PstI - SbfI (58) SphI (64) HindIII (66) KasI (230) NarI (231) SfoI (232) PluTI (234) BseRI (509) BsrGI (729) PfoI (975) BtgI (988) EcoRV (1103) BstXI (1142) BbsI (1397) AflII (1534) AgeI (1599) BspDI - ClaI (1699) BstEII (1814) EcoO109I - PpuMI (1959) YIp351 4406 bp
Eco53kI  (23)
1 site
G A G C T C C T C G A G
BspQI  (4182)
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  (4182)
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.
DrdI  (3963)
1 site
G A C N N N N N N G T C C T G N N N N N N C A G

Sticky ends from different DrdI sites may not be compatible.
PspFI  (3765)
1 site
C C C A G C G G G T C G
BseYI  (3761)
1 site
C C C A G C G G G T C G

After cleavage, BseYI can remain bound to DNA and alter its electrophoretic mobility.
AlwNI  (3656)
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  (3177)
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.
BsaI  (3111)
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.
BpmI  (3108)
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.
NmeAIII  (3030)
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).
FspI  (2954)
1 site
T G C G C A A C G C G T
ScaI  (2696)
1 site
A G T A C T T C A T G A
BanII  (25)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
SacI  (25)
1 site
G A G C T C C T C G A G
AvaI  (31)
1 site
C Y C G R G G R G C Y C

Sticky ends from different AvaI sites may not be compatible.
BsoBI  (31)
1 site
C Y C G R G G R G C Y C

Sticky ends from different BsoBI sites may not be compatible.
BsoBI is typically used at 37°C, but can be used at temperatures up to 65°C.
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
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.
BseRI  (509)
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.
BsrGI  (729)
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.
PfoI  (975)
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.
BtgI  (988)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
EcoRV  (1103)
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.
BstXI  (1142)
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.
BbsI  (1397)
1 site
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
AflII  (1534)
1 site
C T T A A G G A A T T C
AgeI  (1599)
1 site
A C C G G T T G G C C A
BspDI  (1699)
1 site
A T C G A T T A G C T A
ClaI  (1699)
1 site
A T C G A T T A G C T A
BstEII  (1814)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
EcoO109I  (1959)
1 site
R G G N C C Y Y C C N G G R

Sticky ends from different EcoO109I sites may not be compatible.
PpuMI  (1959)
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.
LEU2
759 .. 1853  =  1095 bp
364 amino acids  =  39.2 kDa
Product: 3-isopropylmalate dehydrogenase, required for leucine biosynthesis
yeast auxotrophic marker
LEU2
759 .. 1853  =  1095 bp
364 amino acids  =  39.2 kDa
Product: 3-isopropylmalate dehydrogenase, required for leucine biosynthesis
yeast auxotrophic marker
AmpR
2390 .. 3250  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   2390 .. 2458  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2390 .. 3250  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   2459 .. 3250  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2390 .. 3250  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
3421 .. 4009  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
ori
3421 .. 4009  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
LEU2 promoter
1854 .. 2258  =  405 bp
LEU2 promoter
1854 .. 2258  =  405 bp
lacZα
1 .. 303  =  303 bp
100 amino acids  =  11.5 kDa
Product: LacZα fragment of β-galactosidase
lacZα
1 .. 303  =  303 bp
100 amino acids  =  11.5 kDa
Product: LacZα fragment of β-galactosidase
AmpR promoter
2285 .. 2389  =  105 bp
AmpR promoter
2285 .. 2389  =  105 bp
lac promoter
4333 .. 4363  =  31 bp
3 segments
   Segment 1:  -35  
   4333 .. 4338  =  6 bp
promoter for the E. coli lac operon
lac promoter
4333 .. 4363  =  31 bp
3 segments
   Segment 2:  
   4339 .. 4356  =  18 bp
promoter for the E. coli lac operon
lac promoter
4333 .. 4363  =  31 bp
3 segments
   Segment 3:  -10  
   4357 .. 4363  =  7 bp
promoter for the E. coli lac operon
lac promoter
4333 .. 4363  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
4371 .. 4387  =  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
4371 .. 4387  =  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
4395 .. 5  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
4395 .. 5  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  1 .. 303  =  303 bp
ORF:  100 amino acids  =  11.5 kDa
ORF:  2390 .. 3250  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  4334 .. 284  =  357 bp
ORF:  118 amino acids  =  13.4 kDa
ORF:  759 .. 1853  =  1095 bp
ORF:  364 amino acids  =  39.2 kDa
ORF:  2854 .. 3120  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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