pHet-Act1-2

Mammalian vector encoding a transcription factor activation domain and a DNA-binding domain that can be assembled by drug-induced heterodimerization.

Sequence Author: Clontech (TaKaRa)

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BglII (8197) AhdI (7297) BstBI (6003) Bpu10I (5984) BssHII (5843) SacII (5602) BstEII (5522) RsrII (5504) BsiWI (5444) PflFI - Tth111I (5430) SfiI (5275) NaeI (4535) NgoMIV (4533) MfeI (4231) BsrGI (96) SnaBI (493) Eco53kI (727) SacI (729) I-PpoI (851) NheI (1085) BmtI (1089) PaeR7I - XhoI (1091) ATG AleI (1414) MluI (1990) PmlI (2356) PflMI (2493) PciI (2530) BmgBI (2583) ATG BclI * (2976) HpaI (4222) pHet-Act1-2 8202 bp
BglII  (8197)
1 site
A G A T C T T C T A G A
AhdI  (7297)
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.
BstBI  (6003)
1 site
T T C G A A A A G C T T
Bpu10I  (5984)
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  (5843)
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.
SacII  (5602)
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.
BstEII  (5522)
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.
RsrII  (5504)
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.
BsiWI  (5444)
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.
PflFI  (5430)
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  (5430)
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.
SfiI  (5275)
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.
NaeI  (4535)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI recognition sequence.
NgoMIV  (4533)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV recognition sequence.
MfeI  (4231)
1 site
C A A T T G G T T A A C
BsrGI  (96)
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.
SnaBI  (493)
1 site
T A C G T A A T G C A T
Eco53kI  (727)
1 site
G A G C T C C T C G A G
SacI  (729)
1 site
G A G C T C C T C G A G
I-PpoI  (851)
1 site
C T C T C T T A A G G T A G C G A G A G A A T T C C A T C G

I-PpoI is a homing endonuclease that can recognize a variety of similar recognition sequences.
NheI  (1085)
1 site
G C T A G C C G A T C G
BmtI  (1089)
1 site
G C T A G C C G A T C G
PaeR7I  (1091)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (1091)
1 site
C T C G A G G A G C T C
AleI  (1414)
1 site
C A C N N N N G T G G T G N N N N C A C
MluI  (1990)
1 site
A C G C G T T G C G C A
PmlI  (2356)
1 site
C A C G T G G T G C A C
PflMI  (2493)
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.
PciI  (2530)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BmgBI  (2583)
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.
BclI  (2976)
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.
HpaI  (4222)
1 site
G T T A A C C A A T T G
ATG
2647 .. 2649  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
2647 .. 2649  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
c-myc NLS
2656 .. 2682  =  27 bp
9 amino acids  =  997.2 Da
Product: nuclear localization signal of human c-myc (Dang and Lee, 1988)
c-myc NLS
2656 .. 2682  =  27 bp
9 amino acids  =  997.2 Da
Product: nuclear localization signal of human c-myc (Dang and Lee, 1988)
ZFHD1
2689 .. 3051  =  363 bp
121 amino acids  =  14.5 kDa
3 segments
   Segment 1:  Zif268  
   2689 .. 2862  =  174 bp
   58 amino acids  =  6.9 kDa
Product: composite human DNA-binding domain composed of domains from the transcription factors Zif268 and Oct-1 (Pomerantz et al., 1995)
ZFHD1
2689 .. 3051  =  363 bp
121 amino acids  =  14.5 kDa
3 segments
   Segment 2:  
   2863 .. 2868  =  6 bp
   2 amino acids  =  132.1 Da
Product: composite human DNA-binding domain composed of domains from the transcription factors Zif268 and Oct-1 (Pomerantz et al., 1995)
ZFHD1
2689 .. 3051  =  363 bp
121 amino acids  =  14.5 kDa
3 segments
   Segment 3:  Oct-1  
   2869 .. 3051  =  183 bp
   61 amino acids  =  7.5 kDa
Product: composite human DNA-binding domain composed of domains from the transcription factors Zif268 and Oct-1 (Pomerantz et al., 1995)
ZFHD1
2689 .. 3051  =  363 bp
121 amino acids  =  14.5 kDa
3 segments
Product: composite human DNA-binding domain composed of domains from the transcription factors Zif268 and Oct-1 (Pomerantz et al., 1995)
FKBP (DmrA)
3058 .. 3378  =  321 bp
107 amino acids  =  11.8 kDa
Product: human FK506-binding protein FKBP12
FKBP (DmrA)
3058 .. 3378  =  321 bp
107 amino acids  =  11.8 kDa
Product: human FK506-binding protein FKBP12
FKBP (DmrA)
3388 .. 3705  =  318 bp
106 amino acids  =  11.8 kDa
Product: human FK506-binding protein FKBP12
FKBP (DmrA)
3388 .. 3705  =  318 bp
106 amino acids  =  11.8 kDa
Product: human FK506-binding protein FKBP12
FKBP (DmrA)
3712 .. 4032  =  321 bp
107 amino acids  =  11.8 kDa
Product: human FK506-binding protein FKBP12
FKBP (DmrA)
3712 .. 4032  =  321 bp
107 amino acids  =  11.8 kDa
Product: human FK506-binding protein FKBP12
ATG
1102 .. 1104  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
1102 .. 1104  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
FRB* (DmrC)
1114 .. 1392  =  279 bp
93 amino acids  =  11.3 kDa
Product: T2098L mutant of FKBP-rapamycin binding domain of human FRAP
binds synthetic ligands such as AP21967
FRB* (DmrC)
1114 .. 1392  =  279 bp
93 amino acids  =  11.3 kDa
Product: T2098L mutant of FKBP-rapamycin binding domain of human FRAP
binds synthetic ligands such as AP21967
p65 activation domain
1399 .. 1971  =  573 bp
191 amino acids  =  19.4 kDa
Product: activation domain from the NFκB transcription factor p65
p65 activation domain
1399 .. 1971  =  573 bp
191 amino acids  =  19.4 kDa
Product: activation domain from the NFκB transcription factor p65
AmpR
6510 .. 7370  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   6510 .. 6578  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
6510 .. 7370  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   6579 .. 7370  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
6510 .. 7370  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
PuroR
5388 .. 5987  =  600 bp
199 amino acids  =  21.5 kDa
Product: puromycin N-acetyltransferase
confers resistance to puromycin
PuroR
5388 .. 5987  =  600 bp
199 amino acids  =  21.5 kDa
Product: puromycin N-acetyltransferase
confers resistance to puromycin
ori
7541 .. 8129  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
7541 .. 8129  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
IRES
2040 .. 2613  =  574 bp
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
IRES
2040 .. 2613  =  574 bp
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
f1 ori
4408 .. 4863  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
4408 .. 4863  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
CMV enhancer
138 .. 517  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
138 .. 517  =  380 bp
human cytomegalovirus immediate early enhancer
SV40 promoter
4980 .. 5337  =  358 bp
SV40 enhancer and early promoter
SV40 promoter
4980 .. 5337  =  358 bp
SV40 enhancer and early promoter
CMV promoter
518 .. 729  =  212 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
518 .. 729  =  212 bp
human cytomegalovirus (CMV) immediate early promoter
chimeric intron
890 .. 1022  =  133 bp
chimera between introns from human β-globin and immunoglobulin heavy chain genes
chimeric intron
890 .. 1022  =  133 bp
chimera between introns from human β-globin and immunoglobulin heavy chain genes
SV40 poly(A) signal
4101 .. 4222  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
4101 .. 4222  =  122 bp
SV40 polyadenylation signal
AmpR promoter
6405 .. 6509  =  105 bp
AmpR promoter
6405 .. 6509  =  105 bp
poly(A) signal
6051 .. 6099  =  49 bp
synthetic polyadenylation signal
poly(A) signal
6051 .. 6099  =  49 bp
synthetic polyadenylation signal
SV40 ori
5188 .. 5323  =  136 bp
SV40 origin of replication
SV40 ori
5188 .. 5323  =  136 bp
SV40 origin of replication
ORF:  1102 .. 1983  =  882 bp
ORF:  293 amino acids  =  31.7 kDa
ORF:  2647 .. 4044  =  1398 bp
ORF:  465 amino acids  =  52.7 kDa
ORF:  1331 .. 1558  =  228 bp
ORF:  75 amino acids  =  8.7 kDa
ORF:  5388 .. 5987  =  600 bp
ORF:  199 amino acids  =  21.5 kDa
ORF:  6510 .. 7370  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  2652 .. 2906  =  255 bp
ORF:  84 amino acids  =  10.2 kDa
ORF:  5253 .. 6038  =  786 bp
ORF:  261 amino acids  =  27.1 kDa
ORF:  6974 .. 7240  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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