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Plasmid Files

pAsRed2

Vector for expressing AsRed2 in bacteria.

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pAsRed2 Sequence and MappAsRed2.dna
Map and Sequence File   
Sequence Author:  Clontech
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 BspQI - SapI (3326) PciI (3209) AhdI (2321) BmrI (2281) BsaI (2255) NmeAIII (2174) FspI (2098) PvuI (1952) TsoI (1923) ScaI (1840) XmnI (1721) PvuII (55) HindIII (234) SphI (244) PstI - SbfI (250) SalI (252) AccI (253) HincII (254) BamHI (264) TspMI - XmaI (269) SmaI (271) Acc65I (273) AgeI (276) KpnI (277) NcoI (287) BbsI (313) BsaBI * (412) BsaAI (504) BsrGI (635) NgoMIV (697) NaeI (699) FseI (701) BsgI (793) AarI (815) BseRI (910) NotI (990) ApoI - EcoRI (1004) SpeI (1068) BsiWI (1078) PspOMI (1083) ApaI (1087) NdeI (1279) BstAPI (1280) AflII (1336) ZraI (1400) AatII (1402) SspI (1516) pAsRed2 3334 bp
BspQI  (3326)
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  (3326)
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.
PciI  (3209)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AhdI  (2321)
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.
BmrI  (2281)
1 site
A C T G G G ( N ) 4 N T G A C C C ( N ) 4

The 1-base overhangs produced by BmrI may be hard to ligate.
Sticky ends from different BmrI sites may not be compatible.
Unlike most restriction enzymes, BmrI can cleave DNA in the
absence of magnesium.
BsaI  (2255)
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.
NmeAIII  (2174)
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  (2098)
1 site
T G C G C A A C G C G T
PvuI  (1952)
1 site
C G A T C G G C T A G C
TsoI  (1923)
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).
ScaI  (1840)
1 site
A G T A C T T C A T G A
XmnI  (1721)
1 site
G A A N N N N T T C C T T N N N N A A G
PvuII  (55)
1 site
C A G C T G G T C G A C
HindIII  (234)
1 site
A A G C T T T T C G A A
SphI  (244)
1 site
G C A T G C C G T A C G
PstI  (250)
1 site
C T G C A G G A C G T C
SbfI  (250)
1 site
C C T G C A G G G G A C G T C C
SalI  (252)
1 site
G T C G A C C A G C T G
AccI  (253)
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.
HincII  (254)
1 site
G T Y R A C C A R Y T G
BamHI  (264)
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.
TspMI  (269)
1 site
C C C G G G G G G C C C
XmaI  (269)
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.
SmaI  (271)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
Acc65I  (273)
1 site
G G T A C C C C A T G G
AgeI  (276)
1 site
A C C G G T T G G C C A

AgeI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
KpnI  (277)
1 site
G G T A C C C C A T G G
NcoI  (287)
1 site
C C A T G G G G T A C C
BbsI  (313)
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.
BsaBI  (412)
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.
BsaAI  (504)
1 site
Y A C G T R R T G C A Y
BsrGI  (635)
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.
NgoMIV  (697)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV
recognition sequence.
NaeI  (699)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI
recognition sequence.
FseI  (701)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
BsgI  (793)
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).
AarI  (815)
1 site
C A C C T G C ( N ) 4 G T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the AarI
recognition sequence.
Sticky ends from different AarI sites may not be compatible.
After cleavage, AarI can remain bound to DNA and alter its
electrophoretic mobility.
BseRI  (910)
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.
NotI  (990)
1 site
G C G G C C G C C G C C G G C G
ApoI  (1004)
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  (1004)
1 site
G A A T T C C T T A A G
SpeI  (1068)
1 site
A C T A G T T G A T C A
BsiWI  (1078)
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.
PspOMI  (1083)
1 site
G G G C C C C C C G G G
ApaI  (1087)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
NdeI  (1279)
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  (1280)
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.
AflII  (1336)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
ZraI  (1400)
1 site
G A C G T C C T G C A G
AatII  (1402)
1 site
G A C G T C C T G C A G
SspI  (1516)
1 site
A A T A T T T T A T A A
AmpR
1534 .. 2394  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   1534 .. 1602  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1534 .. 2394  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1603 .. 2394  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1534 .. 2394  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AsRed2
289 .. 987  =  699 bp
232 amino acids  =  25.8 kDa
Product: Anemonia sulcata red fluorescent protein
mammalian codon-optimized
AsRed2
289 .. 987  =  699 bp
232 amino acids  =  25.8 kDa
Product: Anemonia sulcata red fluorescent protein
mammalian codon-optimized
ori
2565 .. 3153  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2565 .. 3153  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
AmpR promoter
1429 .. 1533  =  105 bp
AmpR promoter
1429 .. 1533  =  105 bp
lac promoter
143 .. 173  =  31 bp
   Segment 1:  -35  
   143 .. 148  =  6 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
   Segment 2:  
   149 .. 166  =  18 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
   Segment 3:  -10  
   167 .. 173  =  7 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
181 .. 197  =  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
181 .. 197  =  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
205 .. 221  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
205 .. 221  =  17 bp
common sequencing primer, one of multiple similar
variants
ATG
217 .. 219  =  3 bp
1 amino acid  =  149.2 Da
Product: lacZ start codon
ATG
217 .. 219  =  3 bp
1 amino acid  =  149.2 Da
Product: lacZ start codon
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