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Plasmid Files

pHcRed1

Vector for expressing HcRed1 in bacteria.

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pHcRed1 Sequence and MappHcRed1.dna
Map and Sequence File   
Sequence Author:  Clontech
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 BspQI - SapI (3314) AflIII - PciI (3197) AhdI (2309) BmrI (2269) BsaI (2243) PvuI (1940) TsoI (1911) XmnI (1709) PvuII (55) M13 rev 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) BtgI - NcoI - StyI (287) EcoNI (304) FspAI (319) BsrGI (329) NgoMIV (397) NaeI (399) BsaAI - PmlI (423) KasI (433) NarI (434) SfoI (435) PluTI (437) PpuMI (491) XcmI (723) BssHII (840) NotI (978) ApoI - EcoRI (992) StuI (1052) SpeI (1056) BsiWI (1066) PspOMI (1071) ApaI (1075) PfoI (1129) NdeI (1267) BstAPI (1268) AflII (1324) ZraI (1388) AatII (1390) SspI (1504) pHcRed1 3322 bp
BspQI  (3314)
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  (3314)
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.
AflIII  (3197)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
PciI  (3197)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AhdI  (2309)
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  (2269)
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  (2243)
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.
PvuI  (1940)
1 site
C G A T C G G C T A G C
TsoI  (1911)
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).
XmnI  (1709)
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
BtgI  (287)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
NcoI  (287)
1 site
C C A T G G G G T A C C
StyI  (287)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
EcoNI  (304)
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.
FspAI  (319)
1 site
R T G C G C A Y Y A C G C G T R
BsrGI  (329)
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  (397)
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  (399)
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.
BsaAI  (423)
1 site
Y A C G T R R T G C A Y
PmlI  (423)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
KasI  (433)
1 site
G G C G C C C C G C G G
NarI  (434)
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  (435)
1 site
G G C G C C C C G C G G
PluTI  (437)
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.
PpuMI  (491)
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.
XcmI  (723)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
BssHII  (840)
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.
NotI  (978)
1 site
G C G G C C G C C G C C G G C G
ApoI  (992)
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  (992)
1 site
G A A T T C C T T A A G
StuI  (1052)
1 site
A G G C C T T C C G G A
SpeI  (1056)
1 site
A C T A G T T G A T C A
BsiWI  (1066)
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  (1071)
1 site
G G G C C C C C C G G G
ApaI  (1075)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PfoI  (1129)
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  (1267)
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  (1268)
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  (1324)
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  (1388)
1 site
G A C G T C C T G C A G
AatII  (1390)
1 site
G A C G T C C T G C A G
SspI  (1504)
1 site
A A T A T T T T A T A A
AmpR
1522 .. 2382  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   1522 .. 1590  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1522 .. 2382  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1591 .. 2382  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1522 .. 2382  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
HcRed1
289 .. 975  =  687 bp
228 amino acids  =  25.8 kDa
Product: far-red fluorescent variant of Heteractis
crispa
chromoprotein
mammalian codon-optimized
HcRed1
289 .. 975  =  687 bp
228 amino acids  =  25.8 kDa
Product: far-red fluorescent variant of Heteractis
crispa
chromoprotein
mammalian codon-optimized
ori
2553 .. 3141  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2553 .. 3141  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
AmpR promoter
1417 .. 1521  =  105 bp
AmpR promoter
1417 .. 1521  =  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: start codon
ATG
217 .. 219  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
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