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Plasmid Files

pSP73

Cloning vector for in vitro transcription using the SP6 and T7 RNA polymerase promoters. Essentially identical to pSP72 except for the orientation of the MCS.

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pSP73 Sequence and MappSP73.dna
Map and Sequence File   
Sequence Author:  Promega
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 EcoRV (14) BglII (6) SP6 promoter NdeI (2381) PfoI (2243) EcoO109I (2186) AatII (2132) ZraI (2130) SspI (2014) XmnI (1809) ScaI (1690) TsoI (1609) PvuI (1580) FspI (1432) NmeAIII (1358) BglI (1330) BpmI (1280) BmrI (1250) BspDI - ClaI (19) ApoI - EcoRI (24) Eco53kI (32) BanII - SacI (34) Acc65I (36) KpnI - TspMI - XmaI (40) SmaI (42) BamHI (45) XbaI (51) SalI (57) AccI (58) PstI - SbfI (67) SphI (73) HindIII (75) PvuII (83) PaeR7I - PspXI - XhoI (87) T7 promoter HpaI (138) BspQI - SapI (201) AflIII - PciI (317) BseYI (621) PspFI (625) AlwNI (733) pSP73 2464 bp
EcoRV  (14)
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.
BglII  (6)
1 site
A G A T C T T C T A G A
NdeI  (2381)
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.
PfoI  (2243)
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.
EcoO109I  (2186)
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.
AatII  (2132)
1 site
G A C G T C C T G C A G
ZraI  (2130)
1 site
G A C G T C C T G C A G
SspI  (2014)
1 site
A A T A T T T T A T A A
XmnI  (1809)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (1690)
1 site
A G T A C T T C A T G A
TsoI  (1609)
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).
PvuI  (1580)
1 site
C G A T C G G C T A G C
FspI  (1432)
1 site
T G C G C A A C G C G T
NmeAIII  (1358)
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).
BglI  (1330)
1 site
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
BpmI  (1280)
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.
BmrI  (1250)
1 site
A C T G G G ( N ) 4 N T G A C C C ( N ) 4

The 1-base overhangs produced by BmrI may be hard to ligate.
Sticky ends from different BmrI sites may not be compatible.
Unlike most restriction enzymes, BmrI can cleave DNA in the
absence of magnesium.
BspDI  (19)
1 site
A T C G A T T A G C T A
ClaI  (19)
1 site
A T C G A T T A G C T A
ApoI  (24)
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  (24)
1 site
G A A T T C C T T A A G
Eco53kI  (32)
1 site
G A G C T C C T C G A G
BanII  (34)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
SacI  (34)
1 site
G A G C T C C T C G A G
Acc65I  (36)
1 site
G G T A C C C C A T G G
KpnI  (40)
1 site
G G T A C C C C A T G G
TspMI  (40)
1 site
C C C G G G G G G C C C
XmaI  (40)
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  (42)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
BamHI  (45)
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.
XbaI  (51)
1 site
T C T A G A A G A T C T
SalI  (57)
1 site
G T C G A C C A G C T G
AccI  (58)
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.
PstI  (67)
1 site
C T G C A G G A C G T C
SbfI  (67)
1 site
C C T G C A G G G G A C G T C C
SphI  (73)
1 site
G C A T G C C G T A C G
HindIII  (75)
1 site
A A G C T T T T C G A A
PvuII  (83)
1 site
C A G C T G G T C G A C
PaeR7I  (87)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (87)
1 site
V C T C G A G B B G A G C T C V
XhoI  (87)
1 site
C T C G A G G A G C T C
HpaI  (138)
1 site
G T T A A C C A A T T G
BspQI  (201)
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  (201)
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  (317)
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  (317)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BseYI  (621)
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.
PspFI  (625)
1 site
C C C A G C G G G T C G
AlwNI  (733)
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.
MCS
6 .. 93  =  88 bp
multiple cloning site
MCS
6 .. 93  =  88 bp
multiple cloning site
AmpR
1137 .. 1997  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1137 .. 1928  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1137 .. 1997  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   1929 .. 1997  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1137 .. 1997  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
378 .. 966  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
378 .. 966  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
AmpR promoter
1998 .. 2102  =  105 bp
AmpR promoter
1998 .. 2102  =  105 bp
T7 promoter
102 .. 120  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
102 .. 120  =  19 bp
promoter for bacteriophage T7 RNA polymerase
SP6 promoter
2448 .. 2  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
SP6 promoter
2448 .. 2  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
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