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Plasmid Files

pFC27K HaloTag® CMV-neo

Flexi® vector with a kanamycin/G418 resistance marker, for mammalian expression of a protein with a cleavable C-terminal HaloTag®.

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pFC27K HaloTag CMV-neo Sequence and MappFC27K HaloTag CMV-neo.dna
Map and Sequence File   
Sequence Author:  Promega
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 ScaI (5487) MluI (5403) BspEI (5190) ApaLI (4740) PciI (4426) BstXI (4191) BstBI (4152) RsrII (3986) NaeI (3972) NgoMIV (3970) PluTI (3473) SfoI (3471) NarI (3470) KasI (3469) PacI (3257) AvrII (3233) SfiI (3186) SexAI * (3000) KpnI (2887) Acc65I (2883) BglII (5522) BsrGI (96) SpeI (152) NdeI (387) SnaBI (493) T7 promoter NheI (1052) BmtI (1056) AsiSI - PvuI - SgfI (1060) AhdI (1436) Eco53kI - EcoICRI (1444) SacI (1446) PaeR7I - XhoI (1447) PflMI (1533) AleI (1591) BstEII (1609) BmgBI (1743) PasI (1773) BclI * (1973) AclI (1988) PshAI (2038) SgrAI (2125) EcoRI (2388) XbaI (2392) SalI (2398) SbfI (2408) BlpI (2473) NotI (2503) AanI - AanI - PsiI (2626) HpaI (2646) MfeI (2655) pFC27K HaloTag® CMV-neo 5527 bp
ScaI  (5487)
1 site
A G T A C T T C A T G A
MluI  (5403)
1 site
A C G C G T T G C G C A
BspEI  (5190)
1 site
T C C G G A A G G C C T
ApaLI  (4740)
1 site
G T G C A C C A C G T G
PciI  (4426)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BstXI  (4191)
1 site
C C A N N N N N N T G G G G T N N N N N N A C C

Sticky ends from different BstXI sites may not be compatible.
BstBI  (4152)
1 site
T T C G A A A A G C T T
RsrII  (3986)
1 site
C G G W C C G G C C W G G C

Efficient cleavage requires at least two copies of the RsrII
recognition sequence.
Sticky ends from different RsrII sites may not be compatible.
For full activity, add fresh DTT.
NaeI  (3972)
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.
NgoMIV  (3970)
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.
PluTI  (3473)
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.
SfoI  (3471)
1 site
G G C G C C C C G C G G
NarI  (3470)
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.
KasI  (3469)
1 site
G G C G C C C C G C G G
PacI  (3257)
1 site
T T A A T T A A A A T T A A T T
AvrII  (3233)
1 site
C C T A G G G G A T C C
SfiI  (3186)
1 site
G G C C N N N N N G G C C C C G G N N N N N C C G G

Efficient cleavage requires at least two copies of the SfiI
recognition sequence.
Sticky ends from different SfiI sites may not be compatible.
SexAI  (3000)
1 site
A C C W G G T T G G W C C A
* Blocked by Dcm methylation.
Sticky ends from different SexAI sites may not be compatible.
KpnI  (2887)
1 site
G G T A C C C C A T G G
Acc65I  (2883)
1 site
G G T A C C C C A T G G
BglII  (5522)
1 site
A G A T C T T C T A G A
BsrGI  (96)
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.
SpeI  (152)
1 site
A C T A G T T G A T C A
NdeI  (387)
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.
SnaBI  (493)
1 site
T A C G T A A T G C A T
NheI  (1052)
1 site
G C T A G C C G A T C G
BmtI  (1056)
1 site
G C T A G C C G A T C G
AsiSI  (1060)
1 site
G C G A T C G C C G C T A G C G
PvuI  (1060)
1 site
C G A T C G G C T A G C
SgfI  (1060)
1 site
G C G A T C G C C G C T A G C G
AhdI  (1436)
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.
Eco53kI  (1444)
1 site
G A G C T C C T C G A G
EcoICRI  (1444)
1 site
G A G C T C C T C G A G
SacI  (1446)
1 site
G A G C T C C T C G A G
PaeR7I  (1447)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (1447)
1 site
C T C G A G G A G C T C
PflMI  (1533)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
AleI  (1591)
1 site
C A C N N N N G T G G T G N N N N C A C
BstEII  (1609)
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.
BmgBI  (1743)
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.
PasI  (1773)
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.
BclI  (1973)
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.
AclI  (1988)
1 site
A A C G T T T T G C A A
PshAI  (2038)
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.
SgrAI  (2125)
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.
EcoRI  (2388)
1 site
G A A T T C C T T A A G
XbaI  (2392)
1 site
T C T A G A A G A T C T
SalI  (2398)
1 site
G T C G A C C A G C T G
SbfI  (2408)
1 site
C C T G C A G G G G A C G T C C
BlpI  (2473)
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.
NotI  (2503)
1 site
G C G G C C G C C G C C G G C G
AanI  (2626)
1 site
T T A T A A A A T A T T
AanI  (2626)
1 site
T T A T A A A A T A T T
PsiI  (2626)
1 site
T T A T A A A A T A T T
HpaI  (2646)
1 site
G T T A A C C A A T T G
MfeI  (2655)
1 site
C A A T T G G T T A A C
HaloTag®
1498 .. 2379  =  882 bp
294 amino acids  =  33.4 kDa
Product: modified bacterial dehalogenase that forms
covalent bonds with chloroalkane derivatives
HaloTag®
1498 .. 2379  =  882 bp
294 amino acids  =  33.4 kDa
Product: modified bacterial dehalogenase that forms
covalent bonds with chloroalkane derivatives
NeoR/KanR
3342 .. 4136  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
3342 .. 4136  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
ori
4487 .. 5075  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
4487 .. 5075  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
CMV enhancer
138 .. 517  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
138 .. 517  =  380 bp
human cytomegalovirus immediate early enhancer
SV40 promoter
2891 .. 3248  =  358 bp
SV40 enhancer and early promoter
SV40 promoter
2891 .. 3248  =  358 bp
SV40 enhancer and early promoter
barnase
1087 .. 1422  =  336 bp
111 amino acids  =  12.5 kDa
Product: ribonuclease from Bacillus
amyloliquefaciens

The barnase gene is lethal in standard bacterial
transformation strains.
barnase
1087 .. 1422  =  336 bp
111 amino acids  =  12.5 kDa
Product: ribonuclease from Bacillus
amyloliquefaciens

The barnase gene is lethal in standard bacterial
transformation strains.
cer region
5191 .. 5474  =  284 bp
ColE1-derived recombination site that helps to
maintain plasmids as monomers
cer region
5191 .. 5474  =  284 bp
ColE1-derived recombination site that helps to
maintain plasmids as monomers
CMV promoter
518 .. 721  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
518 .. 721  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
chimeric intron
857 .. 989  =  133 bp
chimera between introns from human β-globin and
immunoglobulin heavy chain genes
chimeric intron
857 .. 989  =  133 bp
chimera between introns from human β-globin and
immunoglobulin heavy chain genes
SV40 poly(A) signal
2525 .. 2646  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2525 .. 2646  =  122 bp
SV40 polyadenylation signal
poly(A) signal
4200 .. 4248  =  49 bp
synthetic polyadenylation signal
poly(A) signal
4200 .. 4248  =  49 bp
synthetic polyadenylation signal
EM7 promoter
3263 .. 3310  =  48 bp
synthetic bacterial promoter
EM7 promoter
3263 .. 3310  =  48 bp
synthetic bacterial promoter
TEV site
1462 .. 1482  =  21 bp
7 amino acids  =  901.0 Da
Product: tobacco etch virus (TEV) protease
recognition and cleavage site
TEV protease recognition site
TEV site
1462 .. 1482  =  21 bp
7 amino acids  =  901.0 Da
Product: tobacco etch virus (TEV) protease
recognition and cleavage site
TEV protease recognition site
T7 promoter
1033 .. 1051  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
1033 .. 1051  =  19 bp
promoter for bacteriophage T7 RNA polymerase
SV40 ori
3099 .. 3234  =  136 bp
SV40 origin of replication
SV40 ori
3099 .. 3234  =  136 bp
SV40 origin of replication
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