pCMV-Tag 2A

Mammalian expression vector for tagging proteins with an N-terminal FLAG epitope. For other reading frames, use pCMV-Tag 2B or pCMV-Tag 2C.

Sequence Author: Agilent Technologies

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PciI (4271) ApaLI (3957) BsaI (3342) PfoI (3128) BstBI (3035) RsrII (2869) BsrDI (2586) PflFI - Tth111I (2471) MscI (2435) PluTI (2356) SfoI (2354) NarI (2353) KasI (2352) StuI (2174) NdeI (240) SnaBI (346) NheI (597) BmtI (601) Eco53kI (653) SacI (655) AleI (661) SacII (662) BstXI (663) NotI (668) ATG FLAG SrfI (709) BamHI (714) PstI (730) EcoRI (732) EcoRV (740) HindIII (744) SalI (759) AccI (760) AbsI - PaeR7I - PspXI - XhoI (765) PspOMI (774) ApaI (778) stop codons PvuI (856) BclI * (1010) MfeI (1103) HpaI (1116) BtsI - BtsαI (1192) MluI (1239) DraIII (1469) SV40 promoter SfiI (2128) BseRI (2171) pCMV-Tag 2A 4323 bp
PciI  (4271)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
ApaLI  (3957)
1 site
G T G C A C C A C G T G
BsaI  (3342)
1 site
G G T C T C N C C A G A G N ( N ) 4

Sticky ends from different BsaI sites may not be compatible.
BsaI can be used between 37°C and 50°C.
PfoI  (3128)
1 site
T C C N G G A A G G N C C T

Sticky ends from different PfoI sites may not be compatible.
BstBI  (3035)
1 site
T T C G A A A A G C T T
RsrII  (2869)
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.
BsrDI  (2586)
1 site
G C A A T G N N C G T T A C

Sticky ends from different BsrDI sites may not be compatible.
PflFI  (2471)
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  (2471)
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.
MscI  (2435)
1 site
T G G C C A A C C G G T
PluTI  (2356)
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  (2354)
1 site
G G C G C C C C G C G G
NarI  (2353)
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  (2352)
1 site
G G C G C C C C G C G G
StuI  (2174)
1 site
A G G C C T T C C G G A
NdeI  (240)
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  (346)
1 site
T A C G T A A T G C A T
NheI  (597)
1 site
G C T A G C C G A T C G
BmtI  (601)
1 site
G C T A G C C G A T C G
Eco53kI  (653)
1 site
G A G C T C C T C G A G
SacI  (655)
1 site
G A G C T C C T C G A G
AleI  (661)
1 site
C A C N N N N G T G G T G N N N N C A C
SacII  (662)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII recognition sequence.
BstXI  (663)
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.
NotI  (668)
1 site
G C G G C C G C C G C C G G C G
SrfI  (709)
1 site
G C C C G G G C C G G G C C C G
BamHI  (714)
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.
PstI  (730)
1 site
C T G C A G G A C G T C
EcoRI  (732)
1 site
G A A T T C C T T A A G
EcoRV  (740)
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.
HindIII  (744)
1 site
A A G C T T T T C G A A
SalI  (759)
1 site
G T C G A C C A G C T G
AccI  (760)
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.
AbsI  (765)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (765)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (765)
1 site
V C T C G A G B B G A G C T C V
XhoI  (765)
1 site
C T C G A G G A G C T C
PspOMI  (774)
1 site
G G G C C C C C C G G G
ApaI  (778)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PvuI  (856)
1 site
C G A T C G G C T A G C
BclI  (1010)
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.
MfeI  (1103)
1 site
C A A T T G G T T A A C
HpaI  (1116)
1 site
G T T A A C C A A T T G
BtsI  (1192)
1 site
G C A G T G N N C G T C A C
BtsαI  (1192)
1 site
G C A G T G N N C G T C A C

Sticky ends from different BtsαI sites may not be compatible.
MluI  (1239)
1 site
A C G C G T T G C G C A
DraIII  (1469)
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.
SfiI  (2128)
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  (2171)
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.
NeoR/KanR
2225 .. 3019  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin)
NeoR/KanR
2225 .. 3019  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin)
ori
3627 .. 4215  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3627 .. 4215  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
1245 .. 1700  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
1245 .. 1700  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
SV40 promoter
1833 .. 2190  =  358 bp
SV40 enhancer and early promoter
SV40 promoter
1833 .. 2190  =  358 bp
SV40 enhancer and early promoter
CMV enhancer
66 .. 370  =  305 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
66 .. 370  =  305 bp
human cytomegalovirus immediate early enhancer
CMV promoter
371 .. 574  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
371 .. 574  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
SV40 poly(A) signal
1117 .. 1238  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1117 .. 1238  =  122 bp
SV40 polyadenylation signal
AmpR promoter
1727 .. 1829  =  103 bp
AmpR promoter
1727 .. 1829  =  103 bp
MCS
706 .. 779  =  74 bp
multiple cloning site
MCS
706 .. 779  =  74 bp
multiple cloning site
HSV TK poly(A) signal
3251 .. 3298  =  48 bp
herpesvirus thymidine kinase polyadenylation signal
HSV TK poly(A) signal
3251 .. 3298  =  48 bp
herpesvirus thymidine kinase polyadenylation signal
ATG
679 .. 681  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
679 .. 681  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
FLAG
682 .. 705  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG epitope tag, followed by an enterokinase cleavage site
FLAG
682 .. 705  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG epitope tag, followed by an enterokinase cleavage site
T3 promoter
620 .. 638  =  19 bp
promoter for bacteriophage T3 RNA polymerase
T3 promoter
620 .. 638  =  19 bp
promoter for bacteriophage T3 RNA polymerase
T7 promoter
825 .. 843  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
825 .. 843  =  19 bp
promoter for bacteriophage T7 RNA polymerase
stop codons
787 .. 797  =  11 bp
stop codons in all three reading frames
stop codons
787 .. 797  =  11 bp
stop codons in all three reading frames
ORF:  3040 .. 3489  =  450 bp
ORF:  149 amino acids  =  16.3 kDa
ORF:  2225 .. 3019  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  2397 .. 2783  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  453 .. 680  =  228 bp
ORF:  75 amino acids  =  8.1 kDa
ORF:  2534 .. 3070  =  537 bp
ORF:  178 amino acids  =  19.9 kDa
ORF:  3245 .. 3478  =  234 bp
ORF:  77 amino acids  =  8.6 kDa
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Individual Sequences & Maps

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