pMCSG79

Transposon-containing vector with a 6xHis-TEV leader, for production of recombinant proteins with the Bac-to-Bac® baculovirus system.

Sequence Author: Midwest Center for Structural Genomics

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NgoMIV (127) AvrII (4455) BclI * (4440) HpaI (4304) MfeI (4291) HindIII (4176) SphI (4168) PaeR7I - XhoI (4158) PstI (4155) XbaI (4143) XcmI (4116) TEV site 6xHis ATG NdeI (4048) BamHI (4032) BstZ17I (3884) AccI (3883) SnaBI (3881) BbsI (3757) BsrGI (3410) PflFI - Tth111I (3228) BsmBI - Esp3I (3182) BseRI (2961) EcoRV (2825) SacII (2768) BstXI (2717) MscI (2709) NaeI (129) BanII (161) BsaHI (836) PvuI (1007) FspI (1153) BpmI (1307) AlwNI (1855) BseYI (1960) PspFI (1964) BspQI - SapI (2381) pMCSG79 4815 bp
NgoMIV  (127)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV recognition sequence.
AvrII  (4455)
1 site
C C T A G G G G A T C C
BclI  (4440)
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.
HpaI  (4304)
1 site
G T T A A C C A A T T G
MfeI  (4291)
1 site
C A A T T G G T T A A C
HindIII  (4176)
1 site
A A G C T T T T C G A A
SphI  (4168)
1 site
G C A T G C C G T A C G
PaeR7I  (4158)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (4158)
1 site
C T C G A G G A G C T C
PstI  (4155)
1 site
C T G C A G G A C G T C
XbaI  (4143)
1 site
T C T A G A A G A T C T
XcmI  (4116)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
NdeI  (4048)
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.
BamHI  (4032)
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.
BstZ17I  (3884)
1 site
G T A T A C C A T A T G
AccI  (3883)
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.
SnaBI  (3881)
1 site
T A C G T A A T G C A T
BbsI  (3757)
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.
BsrGI  (3410)
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.
PflFI  (3228)
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  (3228)
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.
BsmBI  (3182)
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.
BsmBI-v2 is an improved version of BsmBI.
Esp3I  (3182)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different Esp3I sites may not be compatible.
BseRI  (2961)
1 site
G A G G A G ( N ) 8 N N C T C C T C ( N ) 8

Sticky ends from different BseRI sites may not be compatible.
BseRI quickly loses activity at 37°C.
Prolonged incubation with BseRI may lead to degradation of the DNA.
EcoRV  (2825)
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.
SacII  (2768)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII recognition sequence.
BstXI  (2717)
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.
MscI  (2709)
1 site
T G G C C A A C C G G T
NaeI  (129)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI recognition sequence.
BanII  (161)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
BsaHI  (836)
1 site
G R C G Y C C Y G C R G

BsaHI is typically used at 37°C, but is even more active at 60°C.
PvuI  (1007)
1 site
C G A T C G G C T A G C
FspI  (1153)
1 site
T G C G C A A C G C G T
BpmI  (1307)
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.
AlwNI  (1855)
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.
BseYI  (1960)
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  (1964)
1 site
C C C A G C G G G T C G
BspQI  (2381)
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  (2381)
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.
AmpR
589 .. 1449  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   589 .. 657  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
589 .. 1449  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   658 .. 1449  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
589 .. 1449  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
1620 .. 2208  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
1620 .. 2208  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
GmR
2802 .. 3335  =  534 bp
177 amino acids  =  19.4 kDa
Product: gentamycin acetyltransferase
confers resistance to gentamycin
GmR
2802 .. 3335  =  534 bp
177 amino acids  =  19.4 kDa
Product: gentamycin acetyltransferase
confers resistance to gentamycin
f1 ori
2 .. 457  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
2 .. 457  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
Tn7R
2511 .. 2735  =  225 bp
mini-Tn7 element (right end of the Tn7 transposon)
Tn7R
2511 .. 2735  =  225 bp
mini-Tn7 element (right end of the Tn7 transposon)
Tn7L
4468 .. 4633  =  166 bp
mini-Tn7 element (left end of the Tn7 transposon)
Tn7L
4468 .. 4633  =  166 bp
mini-Tn7 element (left end of the Tn7 transposon)
SV40 poly(A) signal
4305 .. 4439  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
4305 .. 4439  =  135 bp
SV40 polyadenylation signal
AmpR promoter
484 .. 588  =  105 bp
AmpR promoter
484 .. 588  =  105 bp
polyhedrin promoter
3904 .. 3995  =  92 bp
promoter for the baculovirus polyhedrin gene
polyhedrin promoter
3904 .. 3995  =  92 bp
promoter for the baculovirus polyhedrin gene
ATG
4050 .. 4052  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
4050 .. 4052  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
6xHis
4053 .. 4070  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
4053 .. 4070  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
TEV site
4095 .. 4115  =  21 bp
7 amino acids  =  900.0 Da
Product: tobacco etch virus (TEV) protease recognition and cleavage site
TEV site
4095 .. 4115  =  21 bp
7 amino acids  =  900.0 Da
Product: tobacco etch virus (TEV) protease recognition and cleavage site
Pc promoter
3524 .. 3552  =  29 bp
3 segments
   Segment 3:  -10  
   3524 .. 3529  =  6 bp
class 1 integron promoter
Pc promoter
3524 .. 3552  =  29 bp
3 segments
   Segment 2:  
   3530 .. 3546  =  17 bp
class 1 integron promoter
Pc promoter
3524 .. 3552  =  29 bp
3 segments
   Segment 1:  -35  
   3547 .. 3552  =  6 bp
class 1 integron promoter
Pc promoter
3524 .. 3552  =  29 bp
3 segments
class 1 integron promoter
ORF:  589 .. 1449  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  2645 .. 2947  =  303 bp
ORF:  100 amino acids  =  11.8 kDa
ORF:  3437 .. 3916  =  480 bp
ORF:  159 amino acids  =  18.5 kDa
ORF:  2940 .. 3272  =  333 bp
ORF:  110 amino acids  =  11.9 kDa
ORF:  3546 .. 3788  =  243 bp
ORF:  80 amino acids  =  9.1 kDa
ORF:  1053 .. 1319  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  2802 .. 3335  =  534 bp
ORF:  177 amino acids  =  19.4 kDa
ORF:  3527 .. 3835  =  309 bp
ORF:  102 amino acids  =  11.4 kDa
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