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Plasmid Files

pTimer

Vector for expressing the Fluorescent Timer in bacteria.

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pTimer.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Clontech (TaKaRa)
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BspQI - SapI (3306) AflIII - PciI (3189) AlwNI (2780) BsaI (2235) BpmI (2232) NmeAIII (2154) PvuI (1932) TsoI (1903) ScaI (1820) XmnI (1701) HindIII (234) BfuAI - BspMI (239) SphI (244) SalI (252) AccI (253) HincII (254) BamHI (264) TspMI - XmaI (269) SmaI (271) Acc65I (273) AgeI (276) KpnI (277) FspAI (340) BstEII (432) BbsI (712) SexAI * (809) BstXI (834) MscI (838) BsgI (871) NotI (970) ApoI - EcoRI (984) SpeI (1048) BsiWI (1058) PspOMI (1063) ApaI - BanII (1067) PfoI (1121) NdeI (1259) BstAPI (1260) AflII (1316) ZraI (1380) AatII (1382) SspI (1496) pTimer 3314 bp
BspQI  (3306)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (3306)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be mixed before removing an aliquot.
AflIII  (3189)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
PciI  (3189)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AlwNI  (2780)
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.
BsaI  (2235)
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.
BpmI  (2232)
1 site
C T G G A G ( N ) 14 N N G A C C T C ( N ) 14

Efficient cleavage requires at least two copies of the BpmI recognition sequence.
Sticky ends from different BpmI sites may not be compatible.
After cleavage, BpmI can remain bound to DNA and alter its electrophoretic mobility.
BpmI quickly loses activity at 37°C.
NmeAIII  (2154)
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).
PvuI  (1932)
1 site
C G A T C G G C T A G C
TsoI  (1903)
1 site
T A R C C A ( N ) 9 N N A T Y G G T ( N ) 9

Sticky ends from different TsoI sites may not be compatible.
After cleavage, TsoI can remain bound to DNA and alter its electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
ScaI  (1820)
1 site
A G T A C T T C A T G A
XmnI  (1701)
1 site
G A A N N N N T T C C T T N N N N A A G
HindIII  (234)
1 site
A A G C T T T T C G A A
BfuAI  (239)
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  (239)
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.
SphI  (244)
1 site
G C A T G C C G T A C G
SalI  (252)
1 site
G T C G A C C A G C T G
AccI  (253)
1 site
G T M K A C C A K M T G

Efficient cleavage with AccI requires ≥13 bp on each side of the recognition sequence.
Sticky ends from different AccI sites may not be compatible.
HincII  (254)
1 site
G T Y R A C C A R Y T G
BamHI  (264)
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.
TspMI  (269)
1 site
C C C G G G G G G C C C
XmaI  (269)
1 site
C C C G G G G G G C C C

Efficient cleavage requires at least two copies of the XmaI recognition sequence.
Full cleavage with XmaI may require a long incubation.
SmaI  (271)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
Acc65I  (273)
1 site
G G T A C C C C A T G G
AgeI  (276)
1 site
A C C G G T T G G C C A
KpnI  (277)
1 site
G G T A C C C C A T G G
FspAI  (340)
1 site
R T G C G C A Y Y A C G C G T R
BstEII  (432)
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.
BbsI  (712)
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.
SexAI  (809)
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.
BstXI  (834)
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.
MscI  (838)
1 site
T G G C C A A C C G G T
BsgI  (871)
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).
NotI  (970)
1 site
G C G G C C G C C G C C G G C G
ApoI  (984)
1 site
R A A T T Y Y T T A A R

ApoI is typically used at 50°C, but is 50% active at 37°C.
EcoRI  (984)
1 site
G A A T T C C T T A A G
SpeI  (1048)
1 site
A C T A G T T G A T C A
BsiWI  (1058)
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.
PspOMI  (1063)
1 site
G G G C C C C C C G G G
ApaI  (1067)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
BanII  (1067)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
PfoI  (1121)
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.
NdeI  (1259)
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.
BstAPI  (1260)
1 site
G C A N N N N N T G C C G T N N N N N A C G

Sticky ends from different BstAPI sites may not be compatible.
AflII  (1316)
1 site
C T T A A G G A A T T C
ZraI  (1380)
1 site
G A C G T C C T G C A G
AatII  (1382)
1 site
G A C G T C C T G C A G
SspI  (1496)
1 site
A A T A T T T T A T A A
AmpR
1514 .. 2374  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   1514 .. 1582  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1514 .. 2374  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1583 .. 2374  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1514 .. 2374  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
Timer
289 .. 969  =  681 bp
226 amino acids  =  26.0 kDa
Product: green-to-red fluorescent timer derivative of DsRed
mammalian codon-optimized
Timer
289 .. 969  =  681 bp
226 amino acids  =  26.0 kDa
Product: green-to-red fluorescent timer derivative of DsRed
mammalian codon-optimized
ori
2545 .. 3133  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2545 .. 3133  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
AmpR promoter
1409 .. 1513  =  105 bp
AmpR promoter
1409 .. 1513  =  105 bp
lac promoter
143 .. 173  =  31 bp
   Segment 1:  -35  
   143 .. 148  =  6 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
   Segment 2:  
   149 .. 166  =  18 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
   Segment 3:  -10  
   167 .. 173  =  7 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
181 .. 197  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lac operator
181 .. 197  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
ATG
217 .. 219  =  3 bp
1 amino acid  =  149.2 Da
Product: lacZ start codon
ATG
217 .. 219  =  3 bp
1 amino acid  =  149.2 Da
Product: lacZ start codon
ORF:  217 .. 969  =  753 bp
ORF:  250 amino acids  =  28.6 kDa
ORF:  1514 .. 2374  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  1978 .. 2244  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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