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Plasmid Files

phRL-null

Promoterless mammalian vector encoding humanized Renilla luciferase.

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phRL-null Sequence and MapphRL-null.dna
Map and Sequence File   
Sequence Author:  Promega
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 Eco53kI (10) PaeR7I - XhoI (5) BglII (1) DrdI (3207) PspFI (3009) BseYI (3005) AlwNI (2900) AhdI (2421) BsrFI (2336) AseI (2246) FspI (2198) PvuI (2052) SacI (12) HindIII (15) NdeI (24) NspI - SphI (32) SpeI (34) KasI (40) NarI (41) SfoI (42) PluTI (44) SalI (47) AccI (48) AflIII - MluI (51) EcoRI (58) TspMI - XmaI (65) SmaI (67) PstI (77) BfuAI - BspMI (91) BbsI (175) AflII (264) T7 promoter NheI (299) BmtI (303) NcoI (307) BstAPI - DraIII (497) PfoI * (589) EcoRV (794) Bsu36I (895) NruI (970) ZraI (1004) PflFI - Tth111I (1005) AatII (1006) PasI (1083) EcoO109I (1154) XbaI (1246) EagI - NotI (1253) PsiI (1376) HpaI (1396) MfeI (1405) BspDI - ClaI (1491) BamHI (1498) SspI (1616) phRL-null 3320 bp
Eco53kI  (10)
1 site
G A G C T C C T C G A G
PaeR7I  (5)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (5)
1 site
C T C G A G G A G C T C
BglII  (1)
1 site
A G A T C T T C T A G A
DrdI  (3207)
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.
PspFI  (3009)
1 site
C C C A G C G G G T C G
BseYI  (3005)
1 site
C C C A G C G G G T C G

After cleavage, BseYI can remain bound to DNA and alter its
electrophoretic mobility.
AlwNI  (2900)
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.
AhdI  (2421)
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.
BsrFI  (2336)
1 site
R C C G G Y Y G G C C R

Efficient cleavage requires at least two copies of the BsrFI
recognition sequence.
After cleavage, BsrFI can remain bound to DNA and alter its
electrophoretic mobility.
AseI  (2246)
1 site
A T T A A T T A A T T A
FspI  (2198)
1 site
T G C G C A A C G C G T
PvuI  (2052)
1 site
C G A T C G G C T A G C
SacI  (12)
1 site
G A G C T C C T C G A G
HindIII  (15)
1 site
A A G C T T T T C G A A
NdeI  (24)
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.
NspI  (32)
1 site
R C A T G Y Y G T A C R
SphI  (32)
1 site
G C A T G C C G T A C G
SpeI  (34)
1 site
A C T A G T T G A T C A
KasI  (40)
1 site
G G C G C C C C G C G G
NarI  (41)
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.
SfoI  (42)
1 site
G G C G C C C C G C G G
PluTI  (44)
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.
SalI  (47)
1 site
G T C G A C C A G C T G
AccI  (48)
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.
AflIII  (51)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
MluI  (51)
1 site
A C G C G T T G C G C A
EcoRI  (58)
1 site
G A A T T C C T T A A G
TspMI  (65)
1 site
C C C G G G G G G C C C
XmaI  (65)
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  (67)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
PstI  (77)
1 site
C T G C A G G A C G T C
BfuAI  (91)
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  (91)
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.
BbsI  (175)
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.
AflII  (264)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
NheI  (299)
1 site
G C T A G C C G A T C G
BmtI  (303)
1 site
G C T A G C C G A T C G
NcoI  (307)
1 site
C C A T G G G G T A C C
BstAPI  (497)
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.
DraIII  (497)
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.
PfoI  (589)
1 site
T C C N G G A A G G N C C T
* Blocked by Dcm methylation.
Sticky ends from different PfoI sites may not be compatible.
EcoRV  (794)
1 site
G A T A T C C T A T A G

EcoRV is reportedly more prone than its isoschizomer Eco32I to
delete a base after cleavage.
Bsu36I  (895)
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.
NruI  (970)
1 site
T C G C G A A G C G C T
ZraI  (1004)
1 site
G A C G T C C T G C A G
PflFI  (1005)
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  (1005)
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.
AatII  (1006)
1 site
G A C G T C C T G C A G
PasI  (1083)
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.
EcoO109I  (1154)
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.
XbaI  (1246)
1 site
T C T A G A A G A T C T
EagI  (1253)
1 site
C G G C C G G C C G G C
NotI  (1253)
1 site
G C G G C C G C C G C C G G C G
PsiI  (1376)
1 site
T T A T A A A A T A T T
HpaI  (1396)
1 site
G T T A A C C A A T T G
MfeI  (1405)
1 site
C A A T T G G T T A A C
BspDI  (1491)
1 site
A T C G A T T A G C T A
ClaI  (1491)
1 site
A T C G A T T A G C T A
BamHI  (1498)
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.
SspI  (1616)
1 site
A A T A T T T T A T A A
hRluc
309 .. 1244  =  936 bp
311 amino acids  =  36.0 kDa
Product: Renilla luciferase
human codon-optimized
hRluc
309 .. 1244  =  936 bp
311 amino acids  =  36.0 kDa
Product: Renilla luciferase
human codon-optimized
AmpR
1634 .. 2494  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   1634 .. 1702  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1634 .. 2494  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1703 .. 2494  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1634 .. 2494  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
2665 .. 3253  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2665 .. 3253  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
chimeric intron
104 .. 236  =  133 bp
chimera between introns from human β-globin and
immunoglobulin heavy chain genes
chimeric intron
104 .. 236  =  133 bp
chimera between introns from human β-globin and
immunoglobulin heavy chain genes
SV40 poly(A) signal
1275 .. 1396  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1275 .. 1396  =  122 bp
SV40 polyadenylation signal
AmpR promoter
1529 .. 1633  =  105 bp
AmpR promoter
1529 .. 1633  =  105 bp
MCS
1 .. 78  =  78 bp
multiple cloning site
MCS
1 .. 78  =  78 bp
multiple cloning site
T7 promoter
281 .. 299  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
281 .. 299  =  19 bp
promoter for bacteriophage T7 RNA polymerase
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