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Plasmid Files

p11

Bacterial vector for expressing a protein with an N-terminal 6xHis-TEV cassette.

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p11 Sequence and Mapp11.dna
Map and Sequence File   
Sequence Author:  Midwest Center for Structural Genomics
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 EcoRI (5702) tet promoter BspDI - ClaI (5678) HindIII (5671) T7 terminator BlpI (5434) SpeI (5427) BamHI (5407) BmtI (5390) NheI (5386) NdeI (5381) TEV Site ATG NcoI (5315) RBS XbaI (5276) lac operator T7 promoter BglII (5210) SgrAI (5169) SphI (5021) EcoNI (4956) MluI (4488) BclI * (4474) BstEII (4306) ApaI (4285) PspOMI (4281) BssHII (4077) HpaI (3986) PshAI (3647) EagI (3420) NruI (3389) BfuAI - BspMI (3305) BsmI (3006) BmeT110I (2935) AvaI - BsoBI (2934) MscI * (2917) FspAI (2907) ZraI (75) AatII (77) SspI (191) ScaI (515) PvuI (627) PstI (754) BsaI (930) AhdI (996) AlwNI (1475) PciI (1884) BspQI - SapI (2001) AccI (2116) BstZ17I (2117) BsaAI (2136) PflFI - Tth111I (2142) Bpu10I (2779) p11 5704 bp
EcoRI  (5702)
1 site
G A A T T C C T T A A G
BspDI  (5678)
1 site
A T C G A T T A G C T A
ClaI  (5678)
1 site
A T C G A T T A G C T A
HindIII  (5671)
1 site
A A G C T T T T C G A A
BlpI  (5434)
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.
SpeI  (5427)
1 site
A C T A G T T G A T C A
BamHI  (5407)
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.
BmtI  (5390)
1 site
G C T A G C C G A T C G
NheI  (5386)
1 site
G C T A G C C G A T C G
NdeI  (5381)
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.
NcoI  (5315)
1 site
C C A T G G G G T A C C
XbaI  (5276)
1 site
T C T A G A A G A T C T
BglII  (5210)
1 site
A G A T C T T C T A G A
SgrAI  (5169)
1 site
C R C C G G Y G G Y G G C C R C

Efficient cleavage requires at least two copies of the SgrAI
recognition sequence.
SphI  (5021)
1 site
G C A T G C C G T A C G
EcoNI  (4956)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
MluI  (4488)
1 site
A C G C G T T G C G C A
BclI  (4474)
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.
BstEII  (4306)
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.
ApaI  (4285)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PspOMI  (4281)
1 site
G G G C C C C C C G G G
BssHII  (4077)
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.
HpaI  (3986)
1 site
G T T A A C C A A T T G
PshAI  (3647)
1 site
G A C N N N N G T C C T G N N N N C A G

PshAI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
EagI  (3420)
1 site
C G G C C G G C C G G C
NruI  (3389)
1 site
T C G C G A A G C G C T
BfuAI  (3305)
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  (3305)
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.
BsmI  (3006)
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.
BmeT110I  (2935)
1 site
C Y C G R G G R G C Y C
AvaI  (2934)
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  (2934)
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.
MscI  (2917)
1 site
T G G C C A A C C G G T
* Blocked by Dcm methylation.
FspAI  (2907)
1 site
R T G C G C A Y Y A C G C G T R
ZraI  (75)
1 site
G A C G T C C T G C A G
AatII  (77)
1 site
G A C G T C C T G C A G
SspI  (191)
1 site
A A T A T T T T A T A A
ScaI  (515)
1 site
A G T A C T T C A T G A
PvuI  (627)
1 site
C G A T C G G C T A G C
PstI  (754)
1 site
C T G C A G G A C G T C
BsaI  (930)
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  (996)
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  (1475)
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  (1884)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (2001)
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  (2001)
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.
AccI  (2116)
1 site
G T M K A C C A K M T G

Efficient cleavage with AccI requires ≥13 bp on each side of the
recognition sequence.
Sticky ends from different AccI sites may not be compatible.
BstZ17I  (2117)
1 site
G T A T A C C A T A T G
BsaAI  (2136)
1 site
Y A C G T R R T G C A Y
PflFI  (2142)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (2142)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
Bpu10I  (2779)
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.
lacI
3761 .. 4843  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
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).
lacI
3761 .. 4843  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
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).
AmpR
209 .. 1069  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   209 .. 277  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
209 .. 1069  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   278 .. 1069  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
209 .. 1069  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
1240 .. 1828  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1240 .. 1828  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
rop
2258 .. 2449  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at
low copy number
rop
2258 .. 2449  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at
low copy number
AmpR promoter
104 .. 208  =  105 bp
AmpR promoter
104 .. 208  =  105 bp
lacI promoter
4844 .. 4921  =  78 bp
lacI promoter
4844 .. 4921  =  78 bp
MCS
5380 .. 5439  =  60 bp
multiple cloning site
MCS
5380 .. 5439  =  60 bp
multiple cloning site
T7 terminator
5445 .. 5492  =  48 bp
transcription terminator for bacteriophage T7 RNA
polymerase
T7 terminator
5445 .. 5492  =  48 bp
transcription terminator for bacteriophage T7 RNA
polymerase
tet promoter
5667 .. 5695  =  29 bp
   Segment 3:  -10  
   5667 .. 5672  =  6 bp
E. coli promoter for tetracycline efflux protein gene
tet promoter
5667 .. 5695  =  29 bp
   Segment 2:  
   5673 .. 5689  =  17 bp
E. coli promoter for tetracycline efflux protein gene
tet promoter
5667 .. 5695  =  29 bp
   Segment 1:  -35  
   5690 .. 5695  =  6 bp
E. coli promoter for tetracycline efflux protein gene
tet promoter
5667 .. 5695  =  29 bp
3 segments
E. coli promoter for tetracycline efflux protein gene
lac operator
5249 .. 5273  =  25 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
5249 .. 5273  =  25 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).
TEV Site
5359 .. 5379  =  21 bp
7 amino acids  =  870.0 Da
Product: tobacco etch virus (TEV) protease
recognition and cleavage site
TEV Site
5359 .. 5379  =  21 bp
7 amino acids  =  870.0 Da
Product: tobacco etch virus (TEV) protease
recognition and cleavage site
T7 promoter
5230 .. 5248  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
5230 .. 5248  =  19 bp
promoter for bacteriophage T7 RNA polymerase
6xHis
5329 .. 5346  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
5329 .. 5346  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
RBS
5304 .. 5309  =  6 bp
ribosome binding site
RBS
5304 .. 5309  =  6 bp
ribosome binding site
ATG
5317 .. 5319  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
5317 .. 5319  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
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