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pLVX-PTuner Green

Lentiviral vector for fusing a destabilization domain to the N-terminus of a partner protein, with co-expression of ZsGreen1.

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pLVX-PTuner Green Sequence and MappLVX-PTuner Green.dna
Map and Sequence File   
Sequence Author:  Clontech
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 SgrDI (8169) SspI (8052) PvuI (7618) FspI (7470) DrdI (6463) BmtI (5972) NheI (5968) StuI (5957) Bsu36I (5178) PstI (5063) PfoI (4731) MluI (4445) BsrGI (4299) BssHII (711) NruI * (833) FseI (1150) MfeI (1189) BbvCI - Bpu10I (1424) BstAPI (1453) AleI (1577) KflI - PpuMI (1934) BspDI - ClaI (2180) CMV enhancer NdeI (2374) SnaBI (2480) SwaI (2797) TspMI - XmaI (2923) SmaI (2925) AgeI (3129) EcoRI (3135) PaeR7I - XhoI (3141) SpeI (3147) MCS XbaI (3153) NotI (3160) BamHI (3167) PspOMI (3288) ApaI (3292) AvrII (3326) BclI * (3831) SgrAI (3836) PflFI - Tth111I (3983) pLVX-PTuner Green 8536 bp
SgrDI  (8169)
1 site
C G T C G A C G G C A G C T G C
SspI  (8052)
1 site
A A T A T T T T A T A A
PvuI  (7618)
1 site
C G A T C G G C T A G C
FspI  (7470)
1 site
T G C G C A A C G C G T
DrdI  (6463)
1 site
G A C N N N N N N G T C C T G N N N N N N C A G

Sticky ends from different DrdI sites may not be compatible.
BmtI  (5972)
1 site
G C T A G C C G A T C G
NheI  (5968)
1 site
G C T A G C C G A T C G
StuI  (5957)
1 site
A G G C C T T C C G G A
Bsu36I  (5178)
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.
PstI  (5063)
1 site
C T G C A G G A C G T C
PfoI  (4731)
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.
MluI  (4445)
1 site
A C G C G T T G C G C A
BsrGI  (4299)
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.
BssHII  (711)
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.
NruI  (833)
1 site
T C G C G A A G C G C T
* Blocked by Dam methylation.
FseI  (1150)
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.
MfeI  (1189)
1 site
C A A T T G G T T A A C
BbvCI  (1424)
1 site
C C T C A G C G G A G T C G
Bpu10I  (1424)
1 site
C C T N A G C G G A N T C G

Efficient cleavage requires at least two copies of the Bpu10I
recognition sequence.
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.
BstAPI  (1453)
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.
AleI  (1577)
1 site
C A C N N N N G T G G T G N N N N C A C
KflI  (1934)
1 site
G G G W C C C C C C W G G G

Sticky ends from different KflI sites may not be compatible.
PpuMI  (1934)
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.
BspDI  (2180)
1 site
A T C G A T T A G C T A
ClaI  (2180)
1 site
A T C G A T T A G C T A
NdeI  (2374)
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  (2480)
1 site
T A C G T A A T G C A T
SwaI  (2797)
1 site
A T T T A A A T T A A A T T T A

SwaI is typically used at 25°C, but is 50% active at 37°C.
TspMI  (2923)
1 site
C C C G G G G G G C C C
XmaI  (2923)
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  (2925)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
AgeI  (3129)
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.
EcoRI  (3135)
1 site
G A A T T C C T T A A G
PaeR7I  (3141)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (3141)
1 site
C T C G A G G A G C T C
SpeI  (3147)
1 site
A C T A G T T G A T C A
XbaI  (3153)
1 site
T C T A G A A G A T C T
NotI  (3160)
1 site
G C G G C C G C C G C C G G C G
BamHI  (3167)
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.
PspOMI  (3288)
1 site
G G G C C C C C C G G G
ApaI  (3292)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
AvrII  (3326)
1 site
C C T A G G G G A T C C
BclI  (3831)
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.
SgrAI  (3836)
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.
PflFI  (3983)
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  (3983)
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
7175 .. 8035  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   7175 .. 7966  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
7175 .. 8035  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   7967 .. 8035  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
7175 .. 8035  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ZsGreen1
3749 .. 4444  =  696 bp
231 amino acids  =  26.1 kDa
Product: Zoanthus green fluorescent protein
mammalian codon-optimized
ZsGreen1
3749 .. 4444  =  696 bp
231 amino acids  =  26.1 kDa
Product: Zoanthus green fluorescent protein
mammalian codon-optimized
5' LTR
1 .. 634  =  634 bp
5' long terminal repeat (LTR) from HIV-1
5' LTR
1 .. 634  =  634 bp
5' long terminal repeat (LTR) from HIV-1
3' LTR
5253 .. 5886  =  634 bp
3' long terminal repeat (LTR) from HIV-1
3' LTR
5253 .. 5886  =  634 bp
3' long terminal repeat (LTR) from HIV-1
WPRE
4458 .. 5046  =  589 bp
woodchuck hepatitis virus posttranscriptional
regulatory element
WPRE
4458 .. 5046  =  589 bp
woodchuck hepatitis virus posttranscriptional
regulatory element
ori
6416 .. 7004  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
6416 .. 7004  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
IRES
3175 .. 3747  =  573 bp
internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)
IRES
3175 .. 3747  =  573 bp
internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)
DD
2805 .. 3128  =  324 bp
108 amino acids  =  11.9 kDa
   Segment 1:  
   2805 .. 2807  =  3 bp
   1 amino acid  =  149.2 Da
Product: destabilization domain that can be
stabilized by Shield1 in the ProteoTuner™ system
L106P mutant of FKBP12
DD
2805 .. 3128  =  324 bp
108 amino acids  =  11.9 kDa
   Segment 2:  
   2808 .. 3128  =  321 bp
   107 amino acids  =  11.8 kDa
Product: destabilization domain that can be
stabilized by Shield1 in the ProteoTuner™ system
L106P mutant of FKBP12
DD
2805 .. 3128  =  324 bp
108 amino acids  =  11.9 kDa
2 segments
Product: destabilization domain that can be
stabilized by Shield1 in the ProteoTuner™ system
L106P mutant of FKBP12
CMV enhancer
2201 .. 2504  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
2201 .. 2504  =  304 bp
human cytomegalovirus immediate early enhancer
RRE
1303 .. 1536  =  234 bp
The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to
the cytoplasm.
RRE
1303 .. 1536  =  234 bp
The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to
the cytoplasm.
CMV promoter
2505 .. 2708  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
2505 .. 2708  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
SV40 poly(A) signal
8188 .. 8322  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
8188 .. 8322  =  135 bp
SV40 polyadenylation signal
HIV-1 Ψ
681 .. 806  =  126 bp
packaging signal of human immunodeficiency virus
type 1
HIV-1 Ψ
681 .. 806  =  126 bp
packaging signal of human immunodeficiency virus
type 1
cPPT/CTS
2028 .. 2144  =  117 bp
central polypurine tract and central termination
sequence of HIV-1 (lacking the first T)
cPPT/CTS
2028 .. 2144  =  117 bp
central polypurine tract and central termination
sequence of HIV-1 (lacking the first T)
AmpR promoter
8036 .. 8140  =  105 bp
AmpR promoter
8036 .. 8140  =  105 bp
MCS
3129 .. 3172  =  44 bp
multiple cloning site
MCS
3129 .. 3172  =  44 bp
multiple cloning site
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