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Plasmid Files

pmirGLO

Vector for quantitatively analyzing microRNA (miRNA) activity by inserting miRNA binding sites downstream of a firefly luciferase gene.

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pmirGLO Sequence and MappmirGLO.dna
Map and Sequence File   
Sequence Author:  Promega
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 AccI (7339) SalI (7338) XbaI (7332) PaeR7I - PspXI - XhoI (7326) BmtI (7324) NheI (7320) SacI (7318) Eco53kI (7316) DraI - PmeI (7309) BbvCI (6356) AleI (6143) BsrGI (6135) MreI - SgrAI (5708) BbsI (5703) ApaI (5675) PspOMI (5671) PpuMI (5513) BsmBI - SpeI (5344) BglII (5090) MluI (4971) PciI (3994) AhdI (3824) SbfI (7348) NotI (93) PsiI (216) MfeI (245) Acc65I (473) KpnI (477) SV40 promoter SfiI (776) AvrII (823) PflMI (877) PfoI * (1169) EcoRV (1374) Bsu36I (1475) NruI (1550) PasI (1663) BssHII (2395) RsrII (2514) BstXI (2719) BspDI - ClaI (2779) BamHI (2792) SnaBI (2886) AseI (3649) pmirGLO 7350 bp
AccI  (7339)
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.
SalI  (7338)
1 site
G T C G A C C A G C T G
XbaI  (7332)
1 site
T C T A G A A G A T C T
PaeR7I  (7326)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (7326)
1 site
V C T C G A G B B G A G C T C V
XhoI  (7326)
1 site
C T C G A G G A G C T C
BmtI  (7324)
1 site
G C T A G C C G A T C G
NheI  (7320)
1 site
G C T A G C C G A T C G
SacI  (7318)
1 site
G A G C T C C T C G A G
Eco53kI  (7316)
1 site
G A G C T C C T C G A G
DraI  (7309)
1 site
T T T A A A A A A T T T
PmeI  (7309)
1 site
G T T T A A A C C A A A T T T G
BbvCI  (6356)
1 site
C C T C A G C G G A G T C G
AleI  (6143)
1 site
C A C N N N N G T G G T G N N N N C A C
BsrGI  (6135)
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.
MreI  (5708)
1 site
C G C C G G C G G C G G C C G C
SgrAI  (5708)
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.
BbsI  (5703)
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.
ApaI  (5675)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PspOMI  (5671)
1 site
G G G C C C C C C G G G
PpuMI  (5513)
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.
BsmBI  (5344)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different BsmBI sites may not be compatible.
SpeI  (5344)
1 site
A C T A G T T G A T C A
BglII  (5090)
1 site
A G A T C T T C T A G A
MluI  (4971)
1 site
A C G C G T T G C G C A
PciI  (3994)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AhdI  (3824)
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.
SbfI  (7348)
1 site
C C T G C A G G G G A C G T C C
NotI  (93)
1 site
G C G G C C G C C G C C G G C G
PsiI  (216)
1 site
T T A T A A A A T A T T
MfeI  (245)
1 site
C A A T T G G T T A A C
Acc65I  (473)
1 site
G G T A C C C C A T G G
KpnI  (477)
1 site
G G T A C C C C A T G G
SfiI  (776)
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.
AvrII  (823)
1 site
C C T A G G G G A T C C
PflMI  (877)
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.
PfoI  (1169)
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  (1374)
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  (1475)
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  (1550)
1 site
T C G C G A A G C G C T
PasI  (1663)
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.
BssHII  (2395)
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.
RsrII  (2514)
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.
BstXI  (2719)
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.
BspDI  (2779)
1 site
A T C G A T T A G C T A
ClaI  (2779)
1 site
A T C G A T T A G C T A
BamHI  (2792)
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.
SnaBI  (2886)
1 site
T A C G T A A T G C A T
AseI  (3649)
1 site
A T T A A T T A A T T A
luciferase
5645 .. 7297  =  1653 bp
550 amino acids  =  60.6 kDa
Product: firefly luciferase
synthetic luc2 version of the luciferase gene
luciferase
5645 .. 7297  =  1653 bp
550 amino acids  =  60.6 kDa
Product: firefly luciferase
synthetic luc2 version of the luciferase gene
hRluc
889 .. 1821  =  933 bp
311 amino acids  =  36.0 kDa
Product: Renilla luciferase
human codon-optimized
hRluc
889 .. 1821  =  933 bp
311 amino acids  =  36.0 kDa
Product: Renilla luciferase
human codon-optimized
AmpR
3037 .. 3897  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   3037 .. 3105  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3037 .. 3897  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   3106 .. 3897  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3037 .. 3897  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
NeoR/KanR
1873 .. 2664  =  792 bp
263 amino acids  =  28.9 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
1873 .. 2664  =  792 bp
263 amino acids  =  28.9 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
ori
4055 .. 4643  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
4055 .. 4643  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
PGK promoter
5099 .. 5598  =  500 bp
mouse phosphoglycerate kinase 1 promoter
PGK promoter
5099 .. 5598  =  500 bp
mouse phosphoglycerate kinase 1 promoter
SV40 promoter
481 .. 838  =  358 bp
SV40 enhancer and early promoter
SV40 promoter
481 .. 838  =  358 bp
SV40 enhancer and early promoter
SV40 poly(A) signal
115 .. 236  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
115 .. 236  =  122 bp
SV40 polyadenylation signal
AmpR promoter
2932 .. 3036  =  105 bp
AmpR promoter
2932 .. 3036  =  105 bp
poly(A) signal
2728 .. 2776  =  49 bp
synthetic polyadenylation signal
poly(A) signal
2728 .. 2776  =  49 bp
synthetic polyadenylation signal
MCS
7306 .. 7350  =  45 bp
multiple cloning site
MCS
7306 .. 7350  =  45 bp
multiple cloning site
SV40 ori
689 .. 824  =  136 bp
SV40 origin of replication
SV40 ori
689 .. 824  =  136 bp
SV40 origin of replication
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