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Plasmid Files

phdKeima-Red-S1

Vector for expressing hdKeima-Red in bacteria.

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phdKeima-Red-S1 Sequence and MapphdKeima-Red-S1.dna
Map and Sequence File   
Sequence Author:  MBL International
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 PfoI (3297) NdeI (3162) HindIII (2945) SphI (2943) PstI (2937) BsaBI * (2903) BtgZI (2890) BmgBI (2854) BsmI (2724) BbsI (2675) PasI (2674) EcoNI (2533) BsaAI (2500) BsrGI (2301) NcoI (2262) BamHI (2253) SmaI (2250) KpnI - TspMI - XmaI (2248) Acc65I (2244) SacI (2242) Eco53kI (2240) ApoI - EcoRI (2232) BspQI - SapI (1993) EcoO109I (8) ZraI (67) AatII (69) SspI (183) XmnI (388) ScaI (507) TsoI (590) NmeAIII (841) BsrFI (903) BsaI (922) AlwNI (1467) phdKeima-Red-S1 3348 bp
PfoI  (3297)
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.
NdeI  (3162)
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.
HindIII  (2945)
1 site
A A G C T T T T C G A A
SphI  (2943)
1 site
G C A T G C C G T A C G
PstI  (2937)
1 site
C T G C A G G A C G T C
BsaBI  (2903)
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.
BtgZI  (2890)
1 site
G C G A T G ( N ) 10 C G C T A C ( N ) 10 ( N ) 4

Sticky ends from different BtgZI sites may not be compatible.
After cleavage, BtgZI can remain bound to DNA and alter its
electrophoretic mobility.
BtgZI is typically used at 60°C, but is 75% active at 37°C.
BmgBI  (2854)
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.
BsmI  (2724)
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.
BbsI  (2675)
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.
PasI  (2674)
1 site
C C C W G G G G G G W C C C

Sticky ends from different PasI sites may not be compatible.
EcoNI  (2533)
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.
BsaAI  (2500)
1 site
Y A C G T R R T G C A Y
BsrGI  (2301)
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.
NcoI  (2262)
1 site
C C A T G G G G T A C C
BamHI  (2253)
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.
SmaI  (2250)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
KpnI  (2248)
1 site
G G T A C C C C A T G G
TspMI  (2248)
1 site
C C C G G G G G G C C C
XmaI  (2248)
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.
Acc65I  (2244)
1 site
G G T A C C C C A T G G
SacI  (2242)
1 site
G A G C T C C T C G A G
Eco53kI  (2240)
1 site
G A G C T C C T C G A G
ApoI  (2232)
1 site
R A A T T Y Y T T A A R

ApoI is typically used at 50°C, but is 50% active at 37°C.
EcoRI  (2232)
1 site
G A A T T C C T T A A G
BspQI  (1993)
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  (1993)
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.
EcoO109I  (8)
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.
ZraI  (67)
1 site
G A C G T C C T G C A G
AatII  (69)
1 site
G A C G T C C T G C A G
SspI  (183)
1 site
A A T A T T T T A T A A
XmnI  (388)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (507)
1 site
A G T A C T T C A T G A
TsoI  (590)
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).
NmeAIII  (841)
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).
BsrFI  (903)
1 site
R C C G G Y Y G G C C R

Efficient cleavage requires at least two copies of the BsrFI
recognition sequence.
After cleavage, BsrFI can remain bound to DNA and alter its
electrophoretic mobility.
BsaI  (922)
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.
AlwNI  (1467)
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.
AmpR
201 .. 1061  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   201 .. 269  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
201 .. 1061  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   270 .. 1061  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
201 .. 1061  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
hdKeima-Red
2264 .. 2932  =  669 bp
222 amino acids  =  25.1 kDa
Product: humanized dimeric Keima-Red fluorescent
protein
mammalian codon-optimized
hdKeima-Red
2264 .. 2932  =  669 bp
222 amino acids  =  25.1 kDa
Product: humanized dimeric Keima-Red fluorescent
protein
mammalian codon-optimized
ori
1232 .. 1820  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1232 .. 1820  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
AmpR promoter
96 .. 200  =  105 bp
AmpR promoter
96 .. 200  =  105 bp
lac promoter
2144 .. 2174  =  31 bp
   Segment 1:  -35  
   2144 .. 2149  =  6 bp
promoter for the E. coli lac operon
lac promoter
2144 .. 2174  =  31 bp
   Segment 2:  
   2150 .. 2167  =  18 bp
promoter for the E. coli lac operon
lac promoter
2144 .. 2174  =  31 bp
   Segment 3:  -10  
   2168 .. 2174  =  7 bp
promoter for the E. coli lac operon
lac promoter
2144 .. 2174  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
2182 .. 2198  =  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
2182 .. 2198  =  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).
M13 rev
2206 .. 2222  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
2206 .. 2222  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
2954 .. 2970  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
2954 .. 2970  =  17 bp
common sequencing primer, one of multiple similar
variants
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