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Plasmid Files

pTK-GLuc

Mammalian vector with a constitutive HSV thymidine kinase promoter for expressing secreted Gaussia luciferase.

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pTK-GLuc Sequence and MappTK-GLuc.dna
Map and Sequence File   
Sequence Author:  New England Biolabs
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 SgrDI (5895) ZraI (5894) SspI (5778) ScaI (5454) TatI (5452) PciI (4081) PfoI (3505) RsrII (3246) AatII (5896) BglII (12) EcoO109I - PpuMI (330) EcoRI (660) MluI (724) HindIII (771) Acc65I (777) KpnI (781) Eco53kI (785) SacI (787) BamHI (789) NruI (907) NotI (1361) AgeI (1368) PaeR7I - XhoI (1374) XbaI (1386) HpaI (1528) MfeI (1537) BspDI - ClaI (1623) DraIII (1996) SexAI * (2287) BseRI (2516) StuI (2519) BclI * (2571) PflFI - Tth111I (2848) pTK-GLuc 5897 bp
SgrDI  (5895)
1 site
C G T C G A C G G C A G C T G C
ZraI  (5894)
1 site
G A C G T C C T G C A G
SspI  (5778)
1 site
A A T A T T T T A T A A
ScaI  (5454)
1 site
A G T A C T T C A T G A
TatI  (5452)
1 site
W G T A C W W C A T G W
PciI  (4081)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PfoI  (3505)
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.
RsrII  (3246)
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.
AatII  (5896)
1 site
G A C G T C C T G C A G
BglII  (12)
1 site
A G A T C T T C T A G A
EcoO109I  (330)
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.
PpuMI  (330)
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.
EcoRI  (660)
1 site
G A A T T C C T T A A G
MluI  (724)
1 site
A C G C G T T G C G C A
HindIII  (771)
1 site
A A G C T T T T C G A A
Acc65I  (777)
1 site
G G T A C C C C A T G G
KpnI  (781)
1 site
G G T A C C C C A T G G
Eco53kI  (785)
1 site
G A G C T C C T C G A G
SacI  (787)
1 site
G A G C T C C T C G A G
BamHI  (789)
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.
NruI  (907)
1 site
T C G C G A A G C G C T
NotI  (1361)
1 site
G C G G C C G C C G C C G G C G
AgeI  (1368)
1 site
A C C G G T T G G C C A

AgeI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
PaeR7I  (1374)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (1374)
1 site
C T C G A G G A G C T C
XbaI  (1386)
1 site
T C T A G A A G A T C T
HpaI  (1528)
1 site
G T T A A C C A A T T G
MfeI  (1537)
1 site
C A A T T G G T T A A C
BspDI  (1623)
1 site
A T C G A T T A G C T A
ClaI  (1623)
1 site
A T C G A T T A G C T A
DraIII  (1996)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
SexAI  (2287)
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.
BseRI  (2516)
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.
StuI  (2519)
1 site
A G G C C T T C C G G A
BclI  (2571)
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.
PflFI  (2848)
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  (2848)
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.
AmpR
4901 .. 5761  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   4901 .. 5692  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4901 .. 5761  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   5693 .. 5761  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4901 .. 5761  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
NeoR/KanR
2602 .. 3396  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
2602 .. 3396  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
HSV TK promoter
19 .. 770  =  752 bp
herpes simplex virus thymidine kinase promoter
HSV TK promoter
19 .. 770  =  752 bp
herpes simplex virus thymidine kinase promoter
ori
4142 .. 4730  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
4142 .. 4730  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
hGLuc
802 .. 1359  =  558 bp
185 amino acids  =  19.9 kDa
Product: secreted Gaussia luciferase
human codon-optimized
hGLuc
802 .. 1359  =  558 bp
185 amino acids  =  19.9 kDa
Product: secreted Gaussia luciferase
human codon-optimized
f1 ori
1763 .. 2191  =  429 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
1763 .. 2191  =  429 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
SV40 promoter
2205 .. 2535  =  331 bp
SV40 enhancer and early promoter
SV40 promoter
2205 .. 2535  =  331 bp
SV40 enhancer and early promoter
SV40 poly(A) signal
1407 .. 1528  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1407 .. 1528  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
3570 .. 3691  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
3570 .. 3691  =  122 bp
SV40 polyadenylation signal
AmpR promoter
5762 .. 5866  =  105 bp
AmpR promoter
5762 .. 5866  =  105 bp
MCS
1360 .. 1391  =  32 bp
multiple cloning site
MCS
1360 .. 1391  =  32 bp
multiple cloning site
SV40 ori
2386 .. 2521  =  136 bp
SV40 origin of replication
SV40 ori
2386 .. 2521  =  136 bp
SV40 origin of replication
Kozak sequence
796 .. 805  =  10 bp
Kozak consensus sequence for GLuc
Kozak sequence
796 .. 805  =  10 bp
Kozak consensus sequence for GLuc
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