pAcGFP1-C In-Fusion Ready

Linearized vector for fusing AcGFP1 to the N-terminus of a partner protein.

Sequence Author: Clontech (TaKaRa)

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PciI (3301) ApaLI (2987) BsaI (2372) BstBI (2065) RsrII (1899) BsrDI (1616) PflFI - Tth111I (1501) FspI (1485) EagI (1289) BspDI * - ClaI * (1223) BseRI (1201) SfiI (1158) CsiI - SexAI * (972) AseI (3366) NdeI (3593) SnaBI (3699) NheI (3950) BmtI (3954) AfeI (3955) AgeI (3959) BstEII (4154) Bpu10I (4164) BssHII (4298) EcoNI (4596) PpuMI (4601) PflMI (4670) BspEI (4689) AflII (4695) End (4711) Start (0) EcoRI (8) BamHI (18) XbaI * (30) BclI * (40) MfeI (133) HincII - HpaI (146) MluI (269) pAcGFP1-C In-Fusion® Ready 4711 bp
PciI  (3301)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
ApaLI  (2987)
1 site
G T G C A C C A C G T G
BsaI  (2372)
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.
BstBI  (2065)
1 site
T T C G A A A A G C T T
RsrII  (1899)
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  (1616)
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  (1501)
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  (1501)
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  (1485)
1 site
T G C G C A A C G C G T
EagI  (1289)
1 site
C G G C C G G C C G G C
BspDI  (1223)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (1223)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
BseRI  (1201)
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.
SfiI  (1158)
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.
CsiI  (972)
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  (972)
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.
AseI  (3366)
1 site
A T T A A T T A A T T A
NdeI  (3593)
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  (3699)
1 site
T A C G T A A T G C A T
NheI  (3950)
1 site
G C T A G C C G A T C G
BmtI  (3954)
1 site
G C T A G C C G A T C G
AfeI  (3955)
1 site
A G C G C T T C G C G A
AgeI  (3959)
1 site
A C C G G T T G G C C A
BstEII  (4154)
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.
Bpu10I  (4164)
1 site
C C T N A G C G G A N T C G

Cleavage may be enhanced when more than one copy of the Bpu10I recognition sequence is present.
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.
BssHII  (4298)
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.
EcoNI  (4596)
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.
PpuMI  (4601)
1 site
R G G W C C Y Y C C W G G R

Sticky ends from different PpuMI sites may not be compatible.
PflMI  (4670)
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.
BspEI  (4689)
1 site
T C C G G A A G G C C T
AflII  (4695)
1 site
C T T A A G G A A T T C
End  (4711)
0 sites
Start  (0)
0 sites
EcoRI  (8)
1 site
G A A T T C C T T A A G
BamHI  (18)
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.
XbaI  (30)
1 site
T C T A G A A G A T C T
* Blocked by Dam methylation.
BclI  (40)
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  (133)
1 site
C A A T T G G T T A A C
HincII  (146)
1 site
G T Y R A C C A R Y T G
HpaI  (146)
1 site
G T T A A C C A A T T G
MluI  (269)
1 site
A C G C G T T G C G C A
NeoR/KanR
1255 .. 2049  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin)
NeoR/KanR
1255 .. 2049  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin)
AcGFP1
3972 .. 4688  =  717 bp
239 amino acids  =  26.9 kDa
Product: Aequorea coerulescens GFP
mammalian codon-optimized
AcGFP1
3972 .. 4688  =  717 bp
239 amino acids  =  26.9 kDa
Product: Aequorea coerulescens GFP
mammalian codon-optimized
ori
2657 .. 3245  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2657 .. 3245  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
275 .. 730  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
275 .. 730  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
SV40 promoter
863 .. 1220  =  358 bp
SV40 enhancer and early promoter
SV40 promoter
863 .. 1220  =  358 bp
SV40 enhancer and early promoter
CMV enhancer
3420 .. 3723  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
3420 .. 3723  =  304 bp
human cytomegalovirus immediate early enhancer
CMV promoter
3724 .. 3927  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
3724 .. 3927  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
SV40 poly(A) signal
147 .. 268  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
147 .. 268  =  122 bp
SV40 polyadenylation signal
AmpR promoter
757 .. 861  =  105 bp
AmpR promoter
757 .. 861  =  105 bp
HSV TK poly(A) signal
2281 .. 2328  =  48 bp
herpesvirus thymidine kinase polyadenylation signal
HSV TK poly(A) signal
2281 .. 2328  =  48 bp
herpesvirus thymidine kinase polyadenylation signal
stop codons
32 .. 42  =  11 bp
stop codons in all three reading frames
stop codons
32 .. 42  =  11 bp
stop codons in all three reading frames
SV40 ori
1071 .. 1206  =  136 bp
SV40 origin of replication
SV40 ori
1071 .. 1206  =  136 bp
SV40 origin of replication
ORF:  1255 .. 2049  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  1427 .. 1813  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  2070 .. 2519  =  450 bp
ORF:  149 amino acids  =  16.3 kDa
ORF:  3972 .. 4709  =  738 bp
ORF:  246 amino acids  =  27.5 kDa
ORF:  1564 .. 2100  =  537 bp
ORF:  178 amino acids  =  19.8 kDa
ORF:  2275 .. 2508  =  234 bp
ORF:  77 amino acids  =  8.6 kDa
ORF:  4188 .. 4709  =  522 bp
ORF:  173 amino acids  =  17.6 kDa  (no start codon)
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