pAAV-AC-RFP

Adeno-associated virus (AAV) vector for expressing a C-terminally TurboRFP-tagged protein using the CMV promoter.

Sequence Author: OriGene

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PciI (5376) NmeAIII (4341) FspI (4265) XmnI (3888) EarI (3689) PfoI (3449) PsiI (3217) PluTI (2789) SfoI (2787) NarI (2786) KasI (2785) CMV enhancer NdeI (390) SnaBI (496) NcoI (516) Eco53kI (722) SacI (724) SacII (811) BsaBI (817) BGI-F sequencing primer (1173 .. 1192) BspDI * - ClaI * (1317) BseRI (1339) AsiSI (1344) MreI - SgrAI (1346) AscI (1350) HindIII (1364) SpeI (1371) MluI (1387) BsiWI (1390) PaeR7I - XhoI (1402) BclI * (1418) BsrGI (1445) RFP-R sequencing primer (1581 .. 1600) PasI (1626) Bsu36I (1661) SfiI (1871) BfuAI - BspMI (1922) PshAI (2035) PmeI (2107) FseI (2117) AleI (2185) BbvCI - Bpu10I (2411) SphI (2444) NsiI (2446) BlpI (2455) BstEII (2616) PmlI (2624) pAAV-AC-RFP 5381 bp
PciI  (5376)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
NmeAIII  (4341)
1 site
G C C G A G ( N ) 18-19 N N C G G C T C ( N ) 18-19

Efficient cleavage requires at least two copies of the NmeAIII recognition sequence.
Sticky ends from different NmeAIII sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
FspI  (4265)
1 site
T G C G C A A C G C G T
XmnI  (3888)
1 site
G A A N N N N T T C C T T N N N N A A G
EarI  (3689)
1 site
C T C T T C N G A G A A G N N N N

Cleavage may be enhanced when more than one copy of the EarI recognition sequence is present.
Sticky ends from different EarI sites may not be compatible.
PfoI  (3449)
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.
PsiI  (3217)
1 site
T T A T A A A A T A T T
PluTI  (2789)
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  (2787)
1 site
G G C G C C C C G C G G
NarI  (2786)
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  (2785)
1 site
G G C G C C C C G C G G
NdeI  (390)
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  (496)
1 site
T A C G T A A T G C A T
NcoI  (516)
1 site
C C A T G G G G T A C C
Eco53kI  (722)
1 site
G A G C T C C T C G A G
SacI  (724)
1 site
G A G C T C C T C G A G
SacII  (811)
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.
BsaBI  (817)
1 site
G A T N N N N A T C C T A N N N N T A G
BspDI  (1317)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (1317)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
BseRI  (1339)
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.
AsiSI  (1344)
1 site
G C G A T C G C C G C T A G C G
MreI  (1346)
1 site
C G C C G G C G G C G G C C G C
SgrAI  (1346)
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  (1350)
1 site
G G C G C G C C C C G C G C G G
HindIII  (1364)
1 site
A A G C T T T T C G A A
SpeI  (1371)
1 site
A C T A G T T G A T C A
MluI  (1387)
1 site
A C G C G T T G C G C A
BsiWI  (1390)
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.
PaeR7I  (1402)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (1402)
1 site
C T C G A G G A G C T C
BclI  (1418)
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.
BsrGI  (1445)
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.
PasI  (1626)
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.
Bsu36I  (1661)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
SfiI  (1871)
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.
BfuAI  (1922)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BfuAI recognition sequence.
Sticky ends from different BfuAI sites may not be compatible.
BfuAI is typically used at 50°C, but is 50% active at 37°C.
BspMI  (1922)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BspMI recognition sequence.
Sticky ends from different BspMI sites may not be compatible.
PshAI  (2035)
1 site
G A C N N N N G T C C T G N N N N C A G

PshAI quickly loses activity at 37°C, but can be used at 25°C for long incubations.
PmeI  (2107)
1 site
G T T T A A A C C A A A T T T G
FseI  (2117)
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.
AleI  (2185)
1 site
C A C N N N N G T G G T G N N N N C A C
BbvCI  (2411)
1 site
C C T C A G C G G A G T C G
Bpu10I  (2411)
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.
SphI  (2444)
1 site
G C A T G C C G T A C G
NsiI  (2446)
1 site
A T G C A T T A C G T A
BlpI  (2455)
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.
BstEII  (2616)
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.
PmlI  (2624)
1 site
C A C G T G G T G C A C
BGI-F sequencing primer
20-mer  /  45% GC
1 binding site
1173 .. 1192  =  20 annealed bases
Tm  =  53°C
RFP-R sequencing primer
20-mer  /  55% GC
1 binding site
1581 .. 1600  =  20 annealed bases
Tm  =  57°C
AmpR
3701 .. 4561  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   3701 .. 3769  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3701 .. 4561  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   3770 .. 4561  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3701 .. 4561  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
TurboRFP
1408 .. 2103  =  696 bp
231 amino acids  =  26.1 kDa
Product: red fluorescent protein from Entacmaea quadricolor
mammalian codon-optimized
TurboRFP
1408 .. 2103  =  696 bp
231 amino acids  =  26.1 kDa
Product: red fluorescent protein from Entacmaea quadricolor
mammalian codon-optimized
ori
4732 .. 5320  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
4732 .. 5320  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
hGH poly(A) signal
2128 .. 2604  =  477 bp
human growth hormone polyadenylation signal
hGH poly(A) signal
2128 .. 2604  =  477 bp
human growth hormone polyadenylation signal
f1 ori
2859 .. 3314  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
2859 .. 3314  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
chimeric intron
873 .. 1251  =  379 bp
fusion between human cytomegalovirus intron A and human β-globin intron 2
chimeric intron
873 .. 1251  =  379 bp
fusion between human cytomegalovirus intron A and human β-globin intron 2
CMV enhancer
217 .. 520  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
217 .. 520  =  304 bp
human cytomegalovirus immediate early enhancer
CMV promoter
521 .. 724  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
521 .. 724  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
AAV2 ITR
1 .. 141  =  141 bp
inverted terminal repeat of adeno-associated virus serotype 2
AAV2 ITR
1 .. 141  =  141 bp
inverted terminal repeat of adeno-associated virus serotype 2
AAV2 ITR
2644 .. 2784  =  141 bp
inverted terminal repeat of adeno-associated virus serotype 2
AAV2 ITR
2644 .. 2784  =  141 bp
inverted terminal repeat of adeno-associated virus serotype 2
AmpR promoter
3596 .. 3700  =  105 bp
AmpR promoter
3596 .. 3700  =  105 bp
MCS
1324 .. 1407  =  84 bp
multiple cloning site
MCS
1324 .. 1407  =  84 bp
multiple cloning site
ORF:  1408 .. 2103  =  696 bp
ORF:  231 amino acids  =  26.1 kDa
ORF:  3701 .. 4561  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  2442 .. 2882  =  441 bp
ORF:  146 amino acids  =  16.0 kDa
ORF:  1509 .. 1754  =  246 bp
ORF:  81 amino acids  =  9.1 kDa
ORF:  2492 .. 2839  =  348 bp
ORF:  115 amino acids  =  13.4 kDa
ORF:  1372 .. 2136  =  765 bp
ORF:  254 amino acids  =  26.3 kDa
ORF:  4165 .. 4431  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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