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p3xFLAG-Myc-CMV™-25

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

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p3xFLAG-Myc-CMV-25.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Sigma-Aldrich
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ScaI (5495) AflIII - PciI (4122) HpaI (3608) MfeI (3595) AfeI (3403) BsrGI (276) SpeI (332) NdeI (567) SnaBI (673) Eco53kI (899) SacI (901) HindIII (1050) NotI (1057) EcoRI (1064) BglII (1076) EcoRV (1084) Acc65I (1088) KpnI (1092) XbaI (1101) BamHI (1140) AleI (1204) EcoO109I (1334) BbvCI - Bpu10I (1430) BlpI (1474) BsmBI (1494) BstXI (1581) AgeI (1692) PflMI (1695) XcmI (1787) SexAI * (1879) SfiI (2066) BseRI (2109) PflFI - Tth111I (2409) BssHII (2688) RsrII (2807) BstBI (2973) NruI (3146) p3xFLAG-Myc-CMV™-25 6406 bp
ScaI  (5495)
1 site
A G T A C T T C A T G A
AflIII  (4122)
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  (4122)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
HpaI  (3608)
1 site
G T T A A C C A A T T G
MfeI  (3595)
1 site
C A A T T G G T T A A C
AfeI  (3403)
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  (1050)
1 site
A A G C T T T T C G A A
NotI  (1057)
1 site
G C G G C C G C C G C C G G C G
EcoRI  (1064)
1 site
G A A T T C C T T A A G
BglII  (1076)
1 site
A G A T C T T C T A G A
EcoRV  (1084)
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  (1088)
1 site
G G T A C C C C A T G G
KpnI  (1092)
1 site
G G T A C C C C A T G G
XbaI  (1101)
1 site
T C T A G A A G A T C T
BamHI  (1140)
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  (1204)
1 site
C A C N N N N G T G G T G N N N N C A C
EcoO109I  (1334)
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  (1430)
1 site
C C T C A G C G G A G T C G
Bpu10I  (1430)
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  (1474)
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  (1494)
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  (1581)
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  (1692)
1 site
A C C G G T T G G C C A
PflMI  (1695)
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  (1787)
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  (1879)
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  (2066)
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  (2109)
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  (2409)
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  (2409)
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  (2688)
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  (2807)
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  (2973)
1 site
T T C G A A A A G C T T
NruI  (3146)
1 site
T C G C G A A G C G C T
AmpR
4942 .. 5802  =  861 bp
286 amino acids  =  31.6