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Plasmid Files

pCMV6-Entry2

Mammalian expression and dual tagging vector that serves as a secondary entry vector to transfer an ORF into a destination vector using the PrecisionShuttle™ system.

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pCMV6-Entry2 Sequence and MappCMV6-Entry2.dna
Map and Sequence File   
Sequence Author:  OriGene
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 BsrGI (301) AanI (5815) DraIII (5690) NruI (5244) EcoNI (4988) AlwNI (4145) ApaLI (4043) PciI (3729) BstZ17I (3350) AanI (3237) BstBI (3060) SpeI (357) CMV enhancer NdeI (592) SnaBI (698) VP1.5 (forward primer) (839 .. 856) Eco53kI (924) SacI (926) T7 promoter EcoRI (979) BamHI (992) Acc65I (998) KpnI (1002) AsiSI - SgfI (1024) MreI - SgrAI (1026) AscI (1030) HindIII (1044) NheI (1055) BmtI (1059) RsrII (1061) MluI (1067) BsiWI (1070) NotI (1076) PaeR7I - PspXI - XhoI (1082) EcoRV (1126) PmeI (1160) FseI (1170) SacII (1174) AleI (1260) XL39 (reverse primer) (1266 .. 1285) AhdI (1316) BbsI (1383) EcoO109I (1390) BsgI (1444) BbvCI (1486) BlpI (1530) BsaI (1592) BstXI (1637) AgeI (1748) XcmI (1843) SexAI * (1935) SfiI (2121) StuI (2167) AvrII (2168) BclI * (2219) BsaBI * (2237) KasI (2377) NarI (2378) SfoI (2379) PluTI (2381) PstI (2431) PflFI - Tth111I (2496) pCMV6-Entry2 5887 bp
BsrGI  (301)
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.
AanI  (5815)
2 sites
T T A T A A A A T A T T
DraIII  (5690)
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.
NruI  (5244)
1 site
T C G C G A A G C G C T
EcoNI  (4988)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
AlwNI  (4145)
1 site
C A G N N N C T G G T C N N N G A C

Sticky ends from different AlwNI sites may not be compatible.
ApaLI  (4043)
1 site
G T G C A C C A C G T G
PciI  (3729)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BstZ17I  (3350)
1 site
G T A T A C C A T A T G
AanI  (3237)
2 sites
T T A T A A A A T A T T
BstBI  (3060)
1 site
T T C G A A A A G C T T
SpeI  (357)
1 site
A C T A G T T G A T C A
NdeI  (592)
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  (698)
1 site
T A C G T A A T G C A T
Eco53kI  (924)
1 site
G A G C T C C T C G A G
SacI  (926)
1 site
G A G C T C C T C G A G
EcoRI  (979)
1 site
G A A T T C C T T A A G
BamHI  (992)
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.
Acc65I  (998)
1 site
G G T A C C C C A T G G
KpnI  (1002)
1 site
G G T A C C C C A T G G
AsiSI  (1024)
1 site
G C G A T C G C C G C T A G C G
SgfI  (1024)
1 site
G C G A T C G C C G C T A G C G
MreI  (1026)
1 site
C G C C G G C G G C G G C C G C
SgrAI  (1026)
1 site
C R C C G G Y G G Y G G C C R C

Efficient cleavage requires at least two copies of the SgrAI
recognition sequence.
AscI  (1030)
1 site
G G C G C G C C C C G C G C G G
HindIII  (1044)
1 site
A A G C T T T T C G A A
NheI  (1055)
1 site
G C T A G C C G A T C G
BmtI  (1059)
1 site
G C T A G C C G A T C G
RsrII  (1061)
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.
MluI  (1067)
1 site
A C G C G T T G C G C A
BsiWI  (1070)
1 site
C G T A C G G C A T G C

BsiWI is typically used at 55°C, but is 50% active at 37°C.
NotI  (1076)
1 site
G C G G C C G C C G C C G G C G
PaeR7I  (1082)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (1082)
1 site
V C T C G A G B B G A G C T C V
XhoI  (1082)
1 site
C T C G A G G A G C T C
EcoRV  (1126)
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.
PmeI  (1160)
1 site
G T T T A A A C C A A A T T T G
FseI  (1170)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
SacII  (1174)
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.
AleI  (1260)
1 site
C A C N N N N G T G G T G N N N N C A C
AhdI  (1316)
1 site
G A C N N N N N G T C C T G N N N N N C A G

The 1-base overhangs produced by AhdI may be hard to ligate.
Sticky ends from different AhdI sites may not be compatible.
BbsI  (1383)
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  (1390)
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  (1444)
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  (1486)
1 site
C C T C A G C G G A G T C G
BlpI  (1530)
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.
BsaI  (1592)
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.
BstXI  (1637)
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  (1748)
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.
XcmI  (1843)
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  (1935)
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  (2121)
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  (2167)
1 site
A G G C C T T C C G G A
AvrII  (2168)
1 site
C C T A G G G G A T C C
BclI  (2219)
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.
BsaBI  (2237)
1 site
G A T N N N N A T C C T A N N N N T A G
* Blocked by Dam methylation.
KasI  (2377)
1 site
G G C G C C C C G C G G
NarI  (2378)
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.
SfoI  (2379)
1 site
G G C G C C C C G C G G
PluTI  (2381)
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.
PstI  (2431)
1 site
C T G C A G G A C G T C
PflFI  (2496)
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  (2496)
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.
VP1.5 (forward primer)
18-mer  /  44% GC
1 binding site
839 .. 856  =  18 annealed bases
Tm  =  51°C
XL39 (reverse primer)
20-mer  /  55% GC
1 binding site
1266 .. 1285  =  20 annealed bases
Tm  =  59°C
KanR
4518 .. 5327  =  810 bp
269 amino acids  =  30.7 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
KanR
4518 .. 5327  =  810 bp
269 amino acids  =  30.7 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
NeoR/KanR
2250 .. 3044  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
2250 .. 3044  =  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
1203 .. 1825  =  623 bp
human growth hormone polyadenylation signal
hGH poly(A) signal
1203 .. 1825  =  623 bp
human growth hormone polyadenylation signal
ori
3790 .. 4375  =  586 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3790 .. 4375  =  586 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
5457 .. 25  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
5457 .. 25  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
CMV enhancer
343 .. 722  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
343 .. 722  =  380 bp
human cytomegalovirus immediate early enhancer
SV40 promoter
1854 .. 2183  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
1854 .. 2183  =  330 bp
SV40 enhancer and early promoter
CMV promoter
723 .. 926  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
723 .. 926  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
SV40 poly(A) signal
3218 .. 3339  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
3218 .. 3339  =  122 bp
SV40 polyadenylation signal
MCS
979 .. 1087  =  109 bp
multiple cloning site
MCS
979 .. 1087  =  109 bp
multiple cloning site
FLAG
1133 .. 1156  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG epitope tag, followed by an
enterokinase cleavage site
FLAG
1133 .. 1156  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG epitope tag, followed by an
enterokinase cleavage site
T7 promoter
952 .. 970  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
952 .. 970  =  19 bp
promoter for bacteriophage T7 RNA polymerase
SV40 ori
2034 .. 2169  =  136 bp
SV40 origin of replication
SV40 ori
2034 .. 2169  =  136 bp
SV40 origin of replication
Myc
1085 .. 1114  =  30 bp
10 amino acids  =  1.2 kDa
Product: Myc (human c-Myc oncogene) epitope tag
Myc
1085 .. 1114  =  30 bp
10 amino acids  =  1.2 kDa
Product: Myc (human c-Myc oncogene) epitope tag
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