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Plasmid Files

pFN19A HaloTag® T7 SP6

Flexi® vector with an ampicillin resistance marker, for cell-free expression of a protein with a cleavable N-terminal HaloTag®.

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pFN19A HaloTag T7 SP6.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Promega
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PflMI (121) BglII (3980) MluI (3861) BspEI (3648) PciI (2884) AgeI (2816) BstBI (2789) AhdI (2714) FspI (2491) AleI (179) BstEII (197) BseRI (199) PflFI - Tth111I (329) BmgBI (331) PasI (361) StuI * (530) BclI * (561) BspDI * - ClaI * (567) PshAI (626) SgrAI (713) BbsI (912) BssHII (938) PaeR7I - XhoI (956) AsiSI - PvuI - SgfI (1014) BtgI - NcoI (1017) DraI - PmeI (1381) Acc65I (1398) KpnI (1402) BamHI (1407) XbaI (1413) PstI - SbfI (1429) BlpI (1494) EcoO109I (1562) SnaBI (1776) SspI (1909) XmnI (2114) pFN19A HaloTag® T7 SP6 4428 bp
PflMI  (121)
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.
BglII  (3980)
1 site
A G A T C T T C T A G A
MluI  (3861)
1 site
A C G C G T T G C G C A
BspEI  (3648)
1 site
T C C G G A A G G C C T
PciI  (2884)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AgeI  (2816)
1 site
A C C G G T T G G C C A
BstBI  (2789)
1 site
T T C G A A A A G C T T
AhdI  (2714)
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.
FspI  (2491)
1 site
T G C G C A A C G C G T
AleI  (179)
1 site
C A C N N N N G T G G T G N N N N C A C
BstEII  (197)
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.
BseRI  (199)
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.
PflFI  (329)
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  (329)
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.
BmgBI  (331)
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  (361)
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.
StuI  (530)
1 site
A G G C C T T C C G G A
* Blocked by Dcm methylation.
BclI  (561)
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.
BspDI  (567)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (567)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
PshAI  (626)
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  (713)
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.
BbsI  (912)
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.
BssHII  (938)
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.
PaeR7I  (956)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (956)
1 site
C T C G A G G A G C T C
AsiSI  (1014)
1 site
G C G A T C G C C G C T A G C G
PvuI  (1014)
1 site
C G A T C G G C T A G C
SgfI  (1014)
1 site
G C G A T C G C C G C T A G C G
BtgI  (1017)
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  (1017)
1 site
C C A T G G G G T A C C
DraI  (1381)
1 site
T T T A A A A A A T T T
PmeI  (1381)
1 site
G T T T A A A C C A A A T T T G
Acc65I  (1398)
1 site
G G T A C C C C A T G G
KpnI  (1402)
1 site
G G T A C C C C A T G G
BamHI  (1407)
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.
XbaI  (1413)
1 site
T C T A G A A G A T C T
PstI  (1429)
1 site
C T G C A G G A C G T C
SbfI  (1429)
1 site
C C T G C A G G G G A C G T C C
BlpI  (1494)
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.
EcoO109I  (1562)
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.
SnaBI  (1776)
1 site
T A C G T A A T G C A T
SspI  (1909)
1 site
A A T A T T T T A T A A
XmnI  (2114)
1 site
G A A N N N N T T C C T T N N N N A A G
HaloTag®
80 .. 970  =  891 bp
297 amino acids  =  33.6 kDa
Product: modified bacterial dehalogenase that forms covalent bonds with chloroalkane derivatives
HaloTag®
80 .. 970  =  891 bp
297 amino acids  =  33.6 kDa
Product: modified bacterial dehalogenase that forms covalent bonds with chloroalkane derivatives
AmpR
1927 .. 2787  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   1927 .. 1995  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1927 .. 2787  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1996 .. 2787  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1927 .. 2787  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
2945 .. 3533  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2945 .. 3533  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
barnase
1041 .. 1376  =  336 bp
111 amino acids  =  12.5 kDa
Product: ribonuclease from Bacillus amyloliquefaciens
The barnase gene is lethal in standard bacterial transformation strains.
barnase
1041 .. 1376  =  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
3649 .. 3932  =  284 bp
ColE1-derived recombination site that helps to maintain plasmids as monomers
cer region
3649 .. 3932  =  284 bp
ColE1-derived recombination site that helps to maintain plasmids as monomers
AmpR promoter
1822 .. 1926  =  105 bp
AmpR promoter
1822 .. 1926  =  105 bp
rrnB T1 terminator
4162 .. 4248  =  87 bp
transcription terminator T1 from the E. coli rrnB gene
rrnB T1 terminator
4162 .. 4248  =  87 bp
transcription terminator T1 from the E. coli rrnB gene
T7 terminator
1546 .. 1593  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 terminator
1546 .. 1593  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
rrnB T2 terminator
4340 .. 4367  =  28 bp
transcription terminator T2 from the E. coli rrnB gene
rrnB T2 terminator
4340 .. 4367  =  28 bp
transcription terminator T2 from the E. coli rrnB gene
TEV site
983 .. 1003  =  21 bp
7 amino acids  =  901.0 Da
Product: tobacco etch virus (TEV) protease recognition and cleavage site
TEV protease recognition site
TEV site
983 .. 1003  =  21 bp
7 amino acids  =  901.0 Da
Product: tobacco etch virus (TEV) protease recognition and cleavage site
TEV protease recognition site
T7 promoter
21 .. 39  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
21 .. 39  =  19 bp
promoter for bacteriophage T7 RNA polymerase
SP6 promoter
45 .. 63  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
SP6 promoter
45 .. 63  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
ORF:  1927 .. 2787  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  80 .. 1027  =  948 bp
ORF:  315 amino acids  =  35.7 kDa
ORF:  1041 .. 1376  =  336 bp
ORF:  111 amino acids  =  12.5 kDa
ORF:  3894 .. 4241  =  348 bp
ORF:  115 amino acids  =  12.4 kDa
ORF:  643 .. 879  =  237 bp
ORF:  78 amino acids  =  8.0 kDa
ORF:  4003 .. 4329  =  327 bp
ORF:  108 amino acids  =  12.1 kDa
ORF:  858 .. 1097  =  240 bp
ORF:  79 amino acids  =  8.9 kDa
ORF:  2391 .. 2657  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  47 .. 1042  =  996 bp
ORF:  331 amino acids  =  37.5 kDa
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