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Plasmid Files

p3xFLAG-Myc-CMV™-26

Vector with a neomycin resistance marker for expression of 3xFLAG- and Myc-tagged proteins in mammalian cells.

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p3xFLAG-Myc-CMV-26 Sequence and Mapp3xFLAG-Myc-CMV-26.dna
Map and Sequence File   
Sequence Author:  Sigma-Aldrich
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 XmnI (5557) ScaI (5438) AflIII - PciI (4065) HpaI (3551) MfeI (3538) AfeI (3346) BsrGI (276) SpeI (332) CMV enhancer NdeI (567) SnaBI (673) Eco53kI (899) SacI (901) ATG HindIII (994) NotI (1001) EcoRI (1008) BglII (1020) EcoRV (1028) Acc65I (1032) KpnI (1036) XbaI (1045) BamHI (1084) AleI (1148) EcoO109I (1278) BsgI (1332) BbvCI - Bpu10I (1374) BlpI (1418) BsmBI (1438) BstXI (1525) AgeI (1636) PflMI (1639) XcmI (1731) SexAI * (1823) SfiI (2009) BseRI (2052) PflFI - Tth111I (2352) BssHII (2631) RsrII (2750) BstBI (2916) NruI (3089) p3xFLAG-Myc-CMV™-26 6332 bp
XmnI  (5557)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (5438)
1 site
A G T A C T T C A T G A
AflIII  (4065)
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  (4065)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
HpaI  (3551)
1 site
G T T A A C C A A T T G
MfeI  (3538)
1 site
C A A T T G G T T A A C
AfeI  (3346)
1 site
A G C G C T T C G C G A
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
HindIII  (994)
1 site
A A G C T T T T C G A A
NotI  (1001)
1 site
G C G G C C G C C G C C G G C G
EcoRI  (1008)
1 site
G A A T T C C T T A A G
BglII  (1020)
1 site
A G A T C T T C T A G A
EcoRV  (1028)
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  (1032)
1 site
G G T A C C C C A T G G
KpnI  (1036)
1 site
G G T A C C C C A T G G
XbaI  (1045)
1 site
T C T A G A A G A T C T
BamHI  (1084)
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.
AleI  (1148)
1 site
C A C N N N N G T G G T G N N N N C A C
EcoO109I  (1278)
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.
BsgI  (1332)
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).
BbvCI  (1374)
1 site
C C T C A G C G G A G T C G
Bpu10I  (1374)
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  (1418)
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  (1438)
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  (1525)
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  (1636)
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  (1639)
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  (1731)
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  (1823)
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  (2009)
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.
BseRI  (2052)
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  (2352)
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  (2352)
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.
BssHII  (2631)
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.
RsrII  (2750)
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.
BstBI  (2916)
1 site
T T C G A A A A G C T T
NruI  (3089)
1 site
T C G C G A A G C G C T
AmpR
4885 .. 5745  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   4885 .. 5676  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4885 .. 5745  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   5677 .. 5745  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4885 .. 5745  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
NeoR/KanR
2106 .. 2900  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin)
NeoR/KanR
2106 .. 2900  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin)
hGH poly(A) signal
1091 .. 1713  =  623 bp
human growth hormone polyadenylation signal
hGH poly(A) signal
1091 .. 1713  =  623 bp
human growth hormone polyadenylation signal
ori
4126 .. 4714  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
4126 .. 4714  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
5877 .. 6332  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
5877 .. 6332  =  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
SV40 promoter
1755 .. 2052  =  298 bp
SV40 enhancer and early promoter
SV40 promoter
1755 .. 2052  =  298 bp
SV40 enhancer and early promoter
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 poly(A) signal
3552 .. 3686  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
3552 .. 3686  =  135 bp
SV40 polyadenylation signal
AmpR promoter
5746 .. 5850  =  105 bp
AmpR promoter
5746 .. 5850  =  105 bp
3xFLAG
931 .. 996  =  66 bp
22 amino acids  =  2.7 kDa
Product: three tandem FLAG epitope tags, followed
by an enterokinase cleavage site
3xFLAG
931 .. 996  =  66 bp
22 amino acids  =  2.7 kDa
Product: three tandem FLAG epitope tags, followed
by an enterokinase cleavage site
Myc
1051 .. 1080  =  30 bp
10 amino acids  =  1.2 kDa
Product: Myc (human c-Myc oncogene) epitope tag
Myc
1051 .. 1080  =  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
3724 .. 3740  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
3724 .. 3740  =  17 bp
common sequencing primer, one of multiple similar
variants
ATG
928 .. 930  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
928 .. 930  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
MCS
994 .. 1050  =  57 bp
multiple cloning site
MCS
994 .. 1050  =  57 bp
multiple cloning site
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