pCMV6-AN-mGFP

PrecisionShuttle™ mammalian expression vector for fusing an ORF to an N-terminal TagGFP2 (mTagGFP) tag.

Sequence Author: OriGene

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AanI (6559) ScaI (5762) PciI (4389) BstZ17I (4010) BsmI (3958) AanI (3897) BstBI (3720) PflFI - Tth111I (3156) CMV enhancer NdeI (592) SnaBI (698) VP1.5 (forward primer) (839 .. 856) Eco53kI (924) SacI (926) EcoRI (979) BamHI (992) Acc65I (998) KpnI (1002) BmgBI (1082) BstEII (1199) PasI (1580) BspEI (1734) AsiSI - SgfI (1750) MreI - SgrAI (1752) AscI (1756) HindIII (1770) RsrII (1787) MluI (1793) NotI (1803) PaeR7I - PspXI - XhoI (1811) PmeI (1820) FseI (1830) XL39 (reverse primer) (1926 .. 1945) BbsI (2043) EcoO109I (2050) BbvCI (2146) BlpI (2190) BsmBI (2210) BstXI (2297) AgeI (2408) SexAI * (2595) SfiI (2781) StuI (2827) AvrII (2828) BclI * (2879) KasI (3037) NarI (3038) SfoI (3039) PluTI (3041) PstI (3091) pCMV6-AN-mGFP 6631 bp
AanI  (6559)
2 sites
T T A T A A A A T A T T
ScaI  (5762)
1 site
A G T A C T T C A T G A
PciI  (4389)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BstZ17I  (4010)
1 site
G T A T A C C A T A T G
BsmI  (3958)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
AanI  (3897)
2 sites
T T A T A A A A T A T T
BstBI  (3720)
1 site
T T C G A A A A G C T T
PflFI  (3156)
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  (3156)
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.
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
BmgBI  (1082)
1 site
C A C G T C G T G C A G

This recognition sequence is asymmetric, so ligating blunt ends generated by BmgBI will not always regenerate a BmgBI site.
BstEII  (1199)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
PasI  (1580)
1 site
C C C W G G G G G G W C C C

Sticky ends from different PasI sites may not be compatible.
BspEI  (1734)
1 site
T C C G G A A G G C C T
AsiSI  (1750)
1 site
G C G A T C G C C G C T A G C G
SgfI  (1750)
1 site
G C G A T C G C C G C T A G C G
MreI  (1752)
1 site
C G C C G G C G G C G G C C G C
SgrAI  (1752)
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  (1756)
1 site
G G C G C G C C C C G C G C G G
HindIII  (1770)
1 site
A A G C T T T T C G A A
RsrII  (1787)
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  (1793)
1 site
A C G C G T T G C G C A
NotI  (1803)
1 site
G C G G C C G C C G C C G G C G
PaeR7I  (1811)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (1811)
1 site
V C T C G A G B B G A G C T C V
XhoI  (1811)
1 site
C T C G A G G A G C T C
PmeI  (1820)
1 site
G T T T A A A C C A A A T T T G
FseI  (1830)
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.
BbsI  (2043)
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  (2050)
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  (2146)
1 site
C C T C A G C G G A G T C G
BlpI  (2190)
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  (2210)
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.
BstXI  (2297)
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  (2408)
1 site
A C C G G T T G G C C A
SexAI  (2595)
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  (2781)
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  (2827)
1 site
A G G C C T T C C G G A
AvrII  (2828)
1 site
C C T A G G G G A T C C
BclI  (2879)
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.
KasI  (3037)
1 site
G G C G C C C C G C G G
NarI  (3038)
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  (3039)
1 site
G G C G C C C C G C G G
PluTI  (3041)
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  (3091)
1 site
C T G C A G G A C G T C
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
1926 .. 1945  =  20 annealed bases
Tm  =  59°C
AmpR
5209 .. 6069  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   5209 .. 6000  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
5209 .. 6069  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   6001 .. 6069  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
5209 .. 6069  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
NeoR/KanR
2910 .. 3704  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
NeoR/KanR
2910 .. 3704  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
TagGFP2
1020 .. 1733  =  714 bp
238 amino acids  =  26.9 kDa
Product: monomeric green fluorescent protein, also known as mTagGFP
mammalian codon-optimized
TagGFP2
1020 .. 1733  =  714 bp
238 amino acids  =  26.9 kDa
Product: monomeric green fluorescent protein, also known as mTagGFP
mammalian codon-optimized
hGH poly(A) signal
1863 .. 2485  =  623 bp
human growth hormone polyadenylation signal
hGH poly(A) signal
1863 .. 2485  =  623 bp
human growth hormone polyadenylation signal
ori
4450 .. 5038  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
4450 .. 5038  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
6201 .. 25  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
6201 .. 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
2514 .. 2843  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
2514 .. 2843  =  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
3878 .. 3999  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
3878 .. 3999  =  122 bp
SV40 polyadenylation signal
AmpR promoter
6070 .. 6174  =  105 bp
AmpR promoter
6070 .. 6174  =  105 bp
MCS
1734 .. 1836  =  103 bp
multiple cloning site
MCS
1734 .. 1836  =  103 bp
multiple cloning 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
2694 .. 2829  =  136 bp
SV40 origin of replication
SV40 ori
2694 .. 2829  =  136 bp
SV40 origin of replication
ORF:  3082 .. 3468  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  5339 .. 5605  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  882 .. 1781  =  900 bp
ORF:  299 amino acids  =  33.8 kDa
ORF:  2910 .. 3704  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  1292 .. 1531  =  240 bp
ORF:  79 amino acids  =  9.4 kDa
ORF:  2227 .. 2451  =  225 bp
ORF:  74 amino acids  =  8.1 kDa
ORF:  5209 .. 6069  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  3219 .. 3755  =  537 bp
ORF:  178 amino acids  =  19.7 kDa
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