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Plasmid Files

pMMB66EH

Broad-host-range bacterial cloning vector. See also pMMB66HE.

 
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pMMB66EH.dna
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TspMI - XmaI (3) EcoRI (8806) MauBI (8531) MluI (8074) ApaI - BanII (7871) PspOMI (7867) HpaI (7572) BsmBI (7459) BspQI - SapI (7266) FspAI (6795) BsaAI (6740) EcoNI (6645) PfoI * (6505) SgrAI (4557) SmaI (5) BamHI (8) SalI (14) PstI (24) HindIII (28) BssS α I (670) DraI (761) PvuI (970) AhdI (1339) SexAI * (1463) SfiI (1697) EcoO109I (2249) BstZ17I (3072) PasI (4078) pMMB66EH 8808 bp
TspMI  (3)
1 site
C C C G G G G G G C C C
XmaI  (3)
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.
EcoRI  (8806)
1 site
G A A T T C C T T A A G
MauBI  (8531)
1 site
C G C G C G C G G C G C G C G C
MluI  (8074)
1 site
A C G C G T T G C G C A
ApaI  (7871)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
BanII  (7871)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
PspOMI  (7867)
1 site
G G G C C C C C C G G G
HpaI  (7572)
1 site
G T T A A C C A A T T G
BsmBI  (7459)
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.
BspQI  (7266)
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  (7266)
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.
FspAI  (6795)
1 site
R T G C G C A Y Y A C G C G T R
BsaAI  (6740)
1 site
Y A C G T R R T G C A Y
EcoNI  (6645)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
PfoI  (6505)
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.
SgrAI  (4557)
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.
SmaI  (5)
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  (8)
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.
SalI  (14)
1 site
G T C G A C C A G C T G
PstI  (24)
1 site
C T G C A G G A C G T C
HindIII  (28)
1 site
A A G C T T T T C G A A
BssSαI  (670)
1 site
C A C G A G G T G C T C
DraI  (761)
1 site
T T T A A A A A A T T T
PvuI  (970)
1 site
C G A T C G G C T A G C
AhdI  (1339)
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.
SexAI  (1463)
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.
SfiI  (1697)
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.
EcoO109I  (2249)
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.
BstZ17I  (3072)
1 site
G T A T A C C A T A T G
PasI  (4078)
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.
lacI
7347 .. 8429  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lacI
7347 .. 8429  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
RSF1010 RepB
3911 .. 4882  =  972 bp
323 amino acids  =  35.9 kDa
Product: replication protein B of the broad-host-range plasmid RSF1010 (Scholz et al., 1989)
RSF1010 RepB
3911 .. 4882  =  972 bp
323 amino acids  =  35.9 kDa
Product: replication protein B of the broad-host-range plasmid RSF1010 (Scholz et al., 1989)
AmpR
552 .. 1412  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   552 .. 620  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
552 .. 1412  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   621 .. 1412  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
552 .. 1412  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
RSF1010 RepC
6219 .. 7070  =  852 bp
283 amino acids  =  31.1 kDa
Product: replication protein C of the broad-host-range plasmid RSF1010 (Scholz et al., 1989)
RSF1010 RepC
6219 .. 7070  =  852 bp
283 amino acids  =  31.1 kDa
Product: replication protein C of the broad-host-range plasmid RSF1010 (Scholz et al., 1989)
RSF1010 oriV
1849 .. 2243  =  395 bp
replication origin of the broad-host-range plasmid RSF1010; requires the RSF1010 RepA/B/C proteins for replication (Scholz et al., 1989)
RSF1010 oriV
1849 .. 2243  =  395 bp
replication origin of the broad-host-range plasmid RSF1010; requires the RSF1010 RepA/B/C proteins for replication (Scholz et al., 1989)
AmpR promoter
460 .. 551  =  92 bp
AmpR promoter
460 .. 551  =  92 bp
RSF1010 oriT
2585 .. 2672  =  88 bp
origin of transfer of the broad-host-range plasmid RSF1010 (Scholz et al., 1989)
RSF1010 oriT
2585 .. 2672  =  88 bp
origin of transfer of the broad-host-range plasmid RSF1010 (Scholz et al., 1989)
rrnB T1 terminator
236 ..