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Plasmid Files

pFLAG-Myc-CMV™-19

Vector for expression and secretion of FLAG- and Myc-tagged proteins in mammalian cells.

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pFLAG-Myc-CMV-19 Sequence and MappFLAG-Myc-CMV-19.dna
Map and Sequence File   
Sequence Author:  Sigma-Aldrich
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 DraIII (4536) NaeI (4430) NgoMIV (4428) ScaI (3864) AflIII - PciI (2491) PsiI (4661) BsrGI (276) SpeI (332) NdeI (567) SnaBI (673) Eco53kI (899) SacI (901) PstI (970) HindIII (998) EagI - NotI (1005) EcoRI (1012) BspDI - ClaI (1019) BglII (1024) EcoRV (1032) Acc65I (1036) KpnI (1040) SalI (1043) AccI (1044) XbaI (1049) BamHI (1055) TspMI - XmaI (1096) SmaI (1098) AleI (1156) BbsI (1279) EcoO109I (1286) BbvCI - Bpu10I (1382) BlpI (1426) BsmBI (1446) BstXI (1533) AgeI (1644) PflMI (1647) XcmI (1739) SexAI * (1831) SfiI (2017) StuI (2063) AvrII (2064) AbsI - PaeR7I - PspXI - XhoI (2086) BspQI - SapI (2375) pFLAG-Myc-CMV™-19 4758 bp
DraIII  (4536)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
NaeI  (4430)
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  (4428)
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.
ScaI  (3864)
1 site
A G T A C T T C A T G A
AflIII  (2491)
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  (2491)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PsiI  (4661)
1 site
T T A T A A A A T A T T
BsrGI  (276)
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  (332)
1 site
A C T A G T T G A T C A
NdeI  (567)
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  (673)
1 site
T A C G T A A T G C A T
Eco53kI  (899)
1 site
G A G C T C C T C G A G
SacI  (901)
1 site
G A G C T C C T C G A G
PstI  (970)
1 site
C T G C A G G A C G T C
HindIII  (998)
1 site
A A G C T T T T C G A A
EagI  (1005)
1 site
C G G C C G G C C G G C
NotI  (1005)
1 site
G C G G C C G C C G C C G G C G
EcoRI  (1012)
1 site
G A A T T C C T T A A G
BspDI  (1019)
1 site
A T C G A T T A G C T A
ClaI  (1019)
1 site
A T C G A T T A G C T A
BglII  (1024)
1 site
A G A T C T T C T A G A
EcoRV  (1032)
1 site
G A T A T C C T A T A G

EcoRV is reportedly more prone than its isoschizomer Eco32I to
delete a base after cleavage.
Acc65I  (1036)
1 site
G G T A C C C C A T G G
KpnI  (1040)
1 site
G G T A C C C C A T G G
SalI  (1043)
1 site
G T C G A C C A G C T G
AccI  (1044)
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.
XbaI  (1049)
1 site
T C T A G A A G A T C T
BamHI  (1055)
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  (1096)
1 site
C C C G G G G G G C C C
XmaI  (1096)
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  (1098)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
AleI  (1156)
1 site
C A C N N N N G T G G T G N N N N C A C
BbsI  (1279)
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.
EcoO109I  (1286)
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.
BbvCI  (1382)
1 site
C C T C A G C G G A G T C G
Bpu10I  (1382)
1 site
C C T N A G C G G A N T C G

Efficient cleavage requires at least two copies of the Bpu10I
recognition sequence.
This recognition sequence is asymmetric, so ligating sticky ends
generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
BlpI  (1426)
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.
BsmBI  (1446)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different BsmBI sites may not be compatible.
BstXI  (1533)
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.
AgeI  (1644)
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.
PflMI  (1647)
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.
XcmI  (1739)
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.
SexAI  (1831)
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.
SfiI  (2017)
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.
StuI  (2063)
1 site
A G G C C T T C C G G A
AvrII  (2064)
1 site
C C T A G G G G A T C C
AbsI  (2086)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (2086)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (2086)
1 site
V C T C G A G B B G A G C T C V
XhoI  (2086)
1 site
C T C G A G G A G C T C
BspQI  (2375)
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  (2375)
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.
AmpR
3311 .. 4171  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   3311 .. 4102  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3311 .. 4171  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   4103 .. 4171  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3311 .. 4171  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
hGH poly(A) signal
1099 .. 1721  =  623 bp
human growth hormone polyadenylation signal
hGH poly(A) signal
1099 .. 1721  =  623 bp
human growth hormone polyadenylation signal
ori
2552 .. 3140  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2552 .. 3140  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
4303 .. 4758  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
4303 .. 4758  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
CMV enhancer
318 .. 697  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
318 .. 697  =  380 bp
human cytomegalovirus immediate early enhancer
CMV promoter
698 .. 901  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
698 .. 901  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
SV40 ori
1930 .. 2065  =  136 bp
SV40 origin of replication
SV40 ori
1930 .. 2065  =  136 bp
SV40 origin of replication
AmpR promoter
4172 .. 4276  =  105 bp
AmpR promoter
4172 .. 4276  =  105 bp
MCS
998 .. 1060  =  63 bp
multiple cloning site
MCS
998 .. 1060  =  63 bp
multiple cloning site
preprotrypsin leader
932 .. 976  =  45 bp
15 amino acids  =  1.4 kDa
Product: leader sequence from mouse preprotrypsin
preprotrypsin leader
932 .. 976  =  45 bp
15 amino acids  =  1.4 kDa
Product: leader sequence from mouse preprotrypsin
Myc
1061 .. 1090  =  30 bp
10 amino acids  =  1.2 kDa
Product: Myc (human c-Myc oncogene) epitope tag
Myc
1061 .. 1090  =  30 bp
10 amino acids  =  1.2 kDa
Product: Myc (human c-Myc oncogene) epitope tag
M13 fwd
141 .. 157  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
141 .. 157  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
2150 .. 2166  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
2150 .. 2166  =  17 bp
common sequencing primer, one of multiple similar
variants
FLAG
977 .. 1000  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG epitope tag, followed by an
enterokinase cleavage site
FLAG
977 .. 1000  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG epitope tag, followed by an
enterokinase cleavage site
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