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Plasmid Files

YEp368

Yeast episomal vector with a LEU2 marker for construction of lacZ fusions. The MCS is reversed in YEp368R.

 
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YEp368.dna
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TspMI - XmaI (17) BspQI - SapI (8366) AhdI (7361) BsaI (7295) ScaI (6880) BsaAI - SnaBI (5846) NsiI (5494) PpuMI (4905) BstEII (4760) AgeI (4545) SmaI (19) BamHI (22) XbaI (28) SalI (34) PstI - SbfI (44) SphI (50) HindIII (52) Bsu36I (279) Bpu10I (560) DraIII (1245) BsaBI * (1386) BclI * (1401) BssHII - MauBI (1553) BstZ17I (2820) BsiWI (2826) NdeI (3012) BlpI (3068) BseRI (3455) BsrGI (3675) AflII (4480) YEp368 8426 bp
TspMI  (17)
1 site
C C C G G G G G G C C C
XmaI  (17)
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.
BspQI  (8366)
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  (8366)
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.
AhdI  (7361)
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  (7295)
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.
ScaI  (6880)
1 site
A G T A C T T C A T G A
BsaAI  (5846)
1 site
Y A C G T R R T G C A Y
SnaBI  (5846)
1 site
T A C G T A A T G C A T
NsiI  (5494)
1 site
A T G C A T T A C G T A
PpuMI  (4905)
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.
BstEII  (4760)
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.
AgeI  (4545)
1 site
A C C G G T T G G C C A
SmaI  (19)
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  (22)
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  (28)
1 site
T C T A G A A G A T C T
SalI  (34)
1 site
G T C G A C C A G C T G
PstI  (44)
1 site
C T G C A G G A C G T C
SbfI  (44)
1 site
C C T G C A G G G G A C G T C C
SphI  (50)
1 site
G C A T G C C G T A C G
HindIII  (52)
1 site
A A G C T T T T C G A A
Bsu36I  (279)
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.
Bpu10I  (560)
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.
DraIII  (1245)
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.
BsaBI  (1386)
1 site
G A T N N N N A T C C T A N N N N T A G
* Blocked by Dam methylation.
BclI  (1401)
1 site
T G A T C A A C T A G T
* Blocked by Dam methylation.
BclI is typically used at 50-55°C, but is 50% active at 37°C.
BssHII  (1553)
1 site
G C G C G C C G C G C G

BssHII is typically used at 50°C, but is 75% active at 37°C.
MauBI  (1553)
1 site
C G C G C G C G G C G C G C G C
BstZ17I  (2820)
1 site
G T A T A C C A T A T G
BsiWI  (2826)
1 site
C G T A C G G C A T G C

BsiWI is typically used at 55°C, but is 50% active at 37°C.
NdeI  (3012)
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.
BlpI  (3068)
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.
BseRI  (3455)
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  (3675)
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.
AflII  (4480)
1 site
C T T A A G G A A T T C
lacZ
68 .. 3115  =  3048 bp
1015 amino acids  =  115.5 kDa
Product: β-galactosidase
lacZ
68 .. 3115  =  3048 bp
1015 amino acids  =  115.5 kDa
Product: β-galactosidase
LEU2
3705 .. 4799  =  1095 bp
364 amino acids  =  39.2 kDa
Product: 3-isopropylmalate dehydrogenase, required for leucine biosynthesis
yeast auxotrophic marker
LEU2
3705 .. 4799  =  1095 bp
364 amino acids  =  39.2 kDa
Product: 3-isopropylmalate dehydrogenase, required for leucine biosynthesis
yeast auxotrophic marker
2μ ori
5205 .. 6187  =  983 bp
yeast 2μ plasmid origin of replication
2μ ori
5205 .. 6187  =  983 bp
yeast 2μ plasmid origin of replication
AmpR
6574 .. 7434  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   6574 .. 6642  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
6574 .. 7434  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   6643 .. 7434  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
6574 .. 7434  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
7605 .. 8193  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
ori
7605 .. 8193  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
LEU2 promoter
4800 .. 5204  =  405 bp
LEU2 promoter
4800 .. 5204  =  405 bp
AmpR promoter
6469 .. 6573  =  105 bp
AmpR promoter
6469 .. 6573  =  105 bp
MCS
1 .. 57  =  57 bp
polylinker
MCS
1 .. 57  =  57 bp
polylinker
ORF:  2989 .. 3249  =  261 bp
ORF:  86 amino acids  =  10.1 kDa
ORF:  6574 .. 7434  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  317 .. 3115  =  2799 bp
ORF:  932 amino acids  =  105.9 kDa
ORF:  5240 .. 5530  =  291 bp
ORF:  96 amino acids  =  11.5 kDa
ORF:  3705 .. 4799  =  1095 bp
ORF:  364 amino acids  =  39.2 kDa
ORF:  7038 .. 7304  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  914 .. 1228  =  315 bp
ORF:  104 amino acids  =  12.0 kDa
ORF:  2339 .. 2575  =  237 bp
ORF:  78 amino acids  =  9.0 kDa
ORF:  1978 .. 2289  =  312 bp
ORF:  103 amino acids  =  10.4 kDa
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