phCMV3

Compact mammalian cell vector for high-level expression of C-terminally HA-tagged proteins.

Sequence Author: MoBiTec

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AflIII - PciI (4193) ApaLI (3879) EcoO109I (3373) BsaI (3264) PfoI (3050) RsrII (2791) BsrDI (2508) PflFI - Tth111I (2393) FspI (2377) MscI (2357) PluTI (2278) SfoI (2276) NarI (2275) KasI (2274) NdeI (234) SnaBI (340) BsmBI - Esp3I (589) BbsI (634) BglII (833) PaeR7I - XhoI (837) HindIII (846) EcoRI (853) Acc65I (869) KpnI (873) PspOMI (877) TspMI - XmaI (880) ApaI (881) SmaI (882) BamHI (884) BspEI (904) XbaI * (929) MfeI (1025) HpaI (1038) BtsI - BtsαI (1114) DraIII (1391) CsiI - SexAI * (1864) SfiI (2050) BseRI (2093) StuI (2096) BspDI * - ClaI * (2115) phCMV3 4251 bp
AflIII  (4193)
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  (4193)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
ApaLI  (3879)
1 site
G T G C A C C A C G T G
EcoO109I  (3373)
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.
BsaI  (3264)
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  (3050)
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.
RsrII  (2791)
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  (2508)
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  (2393)
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  (2393)
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.
FspI  (2377)
1 site
T G C G C A A C G C G T
MscI  (2357)
1 site
T G G C C A A C C G G T
PluTI  (2278)
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  (2276)
1 site
G G C G C C C C G C G G
NarI  (2275)
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  (2274)
1 site
G G C G C C C C G C G G
NdeI  (234)
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  (340)
1 site
T A C G T A A T G C A T
BsmBI  (589)
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.
BsmBI-v2 is an improved version of BsmBI.
Esp3I  (589)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different Esp3I sites may not be compatible.
BbsI  (634)
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.
BglII  (833)
1 site
A G A T C T T C T A G A
PaeR7I  (837)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (837)
1 site
C T C G A G G A G C T C
HindIII  (846)
1 site
A A G C T T T T C G A A
EcoRI  (853)
1 site
G A A T T C C T T A A G
Acc65I  (869)
1 site
G G T A C C C C A T G G
KpnI  (873)
1 site
G G T A C C C C A T G G
PspOMI  (877)
1 site
G G G C C C C C C G G G
TspMI  (880)
1 site
C C C G G G G G G C C C
XmaI  (880)
1 site
C C C G G G G G G C C C

Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
ApaI  (881)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
SmaI  (882)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
BamHI  (884)
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.
BspEI  (904)
1 site
T C C G G A A G G C C T
XbaI  (929)
1 site
T C T A G A A G A T C T
* Blocked by Dam methylation.
MfeI  (1025)
1 site
C A A T T G G T T A A C
HpaI  (1038)
1 site
G T T A A C C A A T T G
BtsI  (1114)
1 site
G C A G T G N N C G T C A C
BtsαI  (1114)
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.
DraIII  (1391)
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.
CsiI  (1864)
1 site
A C C W G G T T G G W C C A

Sticky ends from different CsiI sites may not be compatible.
SexAI  (1864)
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  (2050)
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  (2093)
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.
StuI  (2096)
1 site
A G G C C T T C C G G A
BspDI  (2115)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (2115)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
NeoR/KanR
2147 .. 2941  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin)
NeoR/KanR
2147 .. 2941  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin)
ori
3549 .. 4137  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3549 .. 4137  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
1167 .. 1622  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
1167 .. 1622  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
SV40 promoter
1755 .. 2112  =  358 bp
SV40 enhancer and early promoter
SV40 promoter
1755 .. 2112  =  358 bp
SV40 enhancer and early promoter
CMV enhancer
60 .. 364  =  305 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
60 .. 364  =  305 bp
human cytomegalovirus immediate early enhancer
CMV promoter
365 .. 568  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
365 .. 568  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
SV40 poly(A) signal
1039 .. 1160  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1039 .. 1160  =  122 bp
SV40 polyadenylation signal
CMV intron
704 .. 813  =  110 bp
modified intron from human cytomegalovirus (CMV)
CMV intron
704 .. 813  =  110 bp
modified intron from human cytomegalovirus (CMV)
AmpR promoter
1649 .. 1753  =  105 bp
AmpR promoter
1649 .. 1753  =  105 bp
MCS
833 .. 889  =  57 bp
multiple cloning site
MCS
833 .. 889  =  57 bp
multiple cloning site
HSV TK poly(A) signal
3173 .. 3220  =  48 bp
herpesvirus thymidine kinase polyadenylation signal
HSV TK poly(A) signal
3173 .. 3220  =  48 bp
herpesvirus thymidine kinase polyadenylation signal
HA
890 .. 916  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
HA
890 .. 916  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
SV40 ori
1963 .. 2098  =  136 bp
SV40 origin of replication
SV40 ori
1963 .. 2098  =  136 bp
SV40 origin of replication
T7 promoter
759 .. 777  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
759 .. 777  =  19 bp
promoter for bacteriophage T7 RNA polymerase
ORF:  2962 .. 3411  =  450 bp
ORF:  149 amino acids  =  16.3 kDa
ORF:  524 .. 919  =  396 bp
ORF:  131 amino acids  =  15.4 kDa
ORF:  2147 .. 2941  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  2319 .. 2705  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  2456 .. 2992  =  537 bp
ORF:  178 amino acids  =  19.8 kDa
ORF:  3167 .. 3400  =  234 bp
ORF:  77 amino acids  =  8.6 kDa
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