pMCSG65

Bacterial vector for expressing a protein with a 6xHis-HA-TEV leader plus a second untagged protein.

Sequence Author: Midwest Center for Structural Genomics

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HomePlasmidsStructural Genomics VectorspMCSG65
EagI - NotI (166) PaeR7I - PspXI - XhoI (158) BlpI (80) StyI (57) T7 terminator ScaI (4674) PvuI (4564) PstI (4439) BsaI (4255) AhdI (4194) AlwNI (3717) PciI (3301) BspQI - SapI (3185) BstZ17I (3072) BsaAI (3053) PflFI - Tth111I (3046) HindIII (173) SalI (179) Eco53kI (188) SacI (190) EcoRI (192) BamHI (198) TspMI - XmaI (232) SmaI (234) RBS SspI (275) TEV site Acc65I (300) KpnI (304) BfuAI - BspMI (312) BglII (334) 6xHis ATG NdeI (370) RBS XbaI (408) lac operator BspDI * - ClaI * (477) SgrAI (519) SphI (675) EcoNI (735) PflMI (782) BstAPI (883) MluI (1200) BclI * (1214) BstEII (1381) PspOMI (1407) ApaI (1411) BssHII (1611) EcoRV (1650) HpaI (1706) PshAI (2045) FspAI (2282) PpuMI (2307) Bpu10I (2407) pMCSG65 5369 bp
EagI  (166)
1 site		 C G G C C G G C C G G C
NotI  (166)
1 site		 G C G G C C G C C G C C G G C G
PaeR7I  (158)
1 site		 C T C G A G G A G C T C
PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (158)
1 site		 V C T C G A G B B G A G C T C V
XhoI  (158)
1 site		 C T C G A G G A G C T C
BlpI  (80)
1 site		 G C T N A G C C G A N T C G
Sticky ends from different BlpI sites may not be compatible.
StyI  (57)
1 site		 C C W W G G G G W W C C
Sticky ends from different StyI sites may not be compatible.
ScaI  (4674)
1 site		 A G T A C T T C A T G A
PvuI  (4564)
1 site		 C G A T C G G C T A G C
PstI  (4439)
1 site		 C T G C A G G A C G T C
BsaI  (4255)
1 site		 G G T C T C N C C A G A G N ( N ) 4
Sticky ends from different BsaI sites may not be compatible.
BsaI can be used between 37°C and 50°C.
AhdI  (4194)
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.
AlwNI  (3717)
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.
PciI  (3301)
1 site		 A C A T G T T G T A C A
PciI is inhibited by nonionic detergents.
BspQI  (3185)
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  (3185)
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.
BstZ17I  (3072)
1 site		 G T A T A C C A T A T G
BsaAI  (3053)
1 site		 Y A C G T R R T G C A Y
PflFI  (3046)
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  (3046)
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.
HindIII  (173)
1 site		 A A G C T T T T C G A A
SalI  (179)
1 site		 G T C G A C C A G C T G
Eco53kI  (188)
1 site		 G A G C T C C T C G A G
SacI  (190)
1 site		 G A G C T C C T C G A G
EcoRI  (192)
1 site		 G A A T T C C T T A A G
BamHI  (198)
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.
TspMI  (232)
1 site		 C C C G G G G G G C C C
XmaI  (232)
1 site		 C C C G G G G G G C C C
Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
SmaI  (234)
1 site		 C C C G G G G G G C C C
SmaI can be used at 37°C for brief incubations.
SspI  (275)
1 site		 A A T A T T T T A T A A
Acc65I  (300)
1 site		 G G T A C C C C A T G G
KpnI  (304)
1 site		 G G T A C C C C A T G G
BfuAI  (312)
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  (312)
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.
BglII  (334)
1 site		 A G A T C T T C T A G A
NdeI  (370)
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.
XbaI  (408)
1 site		 T C T A G A A G A T C T
BspDI  (477)
1 site		 A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (477)
1 site		 A T C G A T T A G C T A
* Blocked by Dam methylation.
SgrAI  (519)
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.
SphI  (675)
1 site		 G C A T G C C G T A C G
EcoNI  (735)
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.
PflMI  (782)
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.
BstAPI  (883)
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.
MluI  (1200)
1 site		 A C G C G T T G C G C A
BclI  (1214)
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.
BstEII  (1381)
1 site		 G G T N A C C C C A N T G G
Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
PspOMI  (1407)
1 site		 G G G C C C C C C G G G
ApaI  (1411)
1 site		 G G G C C C C C C G G G
ApaI can be used between 25°C and 37°C.
BssHII  (1611)
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.
EcoRV  (1650)
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.
HpaI  (1706)
1 site		 G T T A A C C A A T T G
PshAI  (2045)
1 site		 G A C N N N N G T C C T G N N N N C A G
PshAI quickly loses activity at 37°C, but can be used at 25°C for long incubations.
FspAI  (2282)
1 site		 R T G C G C A Y Y A C G C G T R
PpuMI  (2307)
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.
Bpu10I  (2407)
1 site		 C C T N A G C G G A N T C G
Cleavage may be enhanced when more than one copy of the Bpu10I recognition sequence is present.
This recognition sequence is asymmetric, so ligating sticky ends generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
lacI
850 .. 1932  =  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
850 .. 1932  =  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).
AmpR
4121 .. 4981  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
   Segment 2:  
   4121 .. 4912  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4121 .. 4981  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
   Segment 1:  signal sequence  
   4913 .. 4981  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4121 .. 4981  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
3362 .. 3950  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3362 .. 3950  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
rop
2741 .. 2932  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at low copy number
rop
2741 .. 2932  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at low copy number
AmpR promoter
4982 .. 5085  =  104 bp
AmpR promoter
4982 .. 5085  =  104 bp
TEV site
279 .. 299  =  21 bp
7 amino acids  =  900.0 Da
Product: tobacco etch virus (TEV) protease recognition and cleavage site
TEV site
279 .. 299  =  21 bp
7 amino acids  =  900.0 Da
Product: tobacco etch virus (TEV) protease recognition and cleavage site
HA
306 .. 332  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
HA
306 .. 332  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
6xHis
351 .. 368  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
351 .. 368  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
ATG
369 .. 371  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
369 .. 371  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
lacI promoter
772 .. 849  =  78 bp
lacI promoter
772 .. 849  =  78 bp
T7 terminator
26 .. 73  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 terminator
26 .. 73  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
lac operator
416 .. 440  =  25 bp
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).
lac operator
416 .. 440  =  25 bp
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).
T7 promoter
441 .. 459  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
441 .. 459  =  19 bp
promoter for bacteriophage T7 RNA polymerase
6xHis
140 .. 157  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
140 .. 157  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
RBS
243 .. 248  =  6 bp
ribosome binding site
RBS
243 .. 248  =  6 bp
ribosome binding site
RBS
380 .. 385  =  6 bp
ribosome binding site
RBS
380 .. 385  =  6 bp
ribosome binding site
ORF:  973 .. 1932  =  960 bp
ORF:  319 amino acids  =  34.1 kDa
ORF:  1988 .. 2344  =  357 bp
ORF:  118 amino acids  =  13.0 kDa
ORF:  2708 .. 2932  =  225 bp
ORF:  74 amino acids  =  8.5 kDa
ORF:  351 .. 1100  =  750 bp
ORF:  249 amino acids  =  26.4 kDa
ORF:  1689 .. 1952  =  264 bp
ORF:  87 amino acids  =  8.9 kDa
ORF:  4251 .. 4517  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  423 .. 674  =  252 bp
ORF:  83 amino acids  =  8.6 kDa
ORF:  548 .. 811  =  264 bp
ORF:  87 amino acids  =  9.5 kDa
ORF:  1715 .. 1966  =  252 bp
ORF:  83 amino acids  =  9.1 kDa
ORF:  4121 .. 4981  =  861 bp
ORF:  286 amino acids  =  31.5 kDa
ORF:  2341 .. 2709  =  369 bp
ORF:  122 amino acids  =  14.2 kDa
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Individual Sequences & Maps

p11
pMCSG19B (linearized)
pMCSG41
pMCSG68 (linearized)
p15Tv-L
pMCSG19C
pMCSG41 (linearized)
pMCSG69
p283
pMCSG19C (linearized)
pMCSG42
pMCSG69 (linearized)
pBEN1-SGC
pMCSG20
pMCSG42 (linearized)
pMCSG7
pBEN1-SGC (linearized)
pMCSG20 (linearized)
pMCSG43
pMCSG7 (linearized)
pDB-HisGST
pMCSG21
pMCSG43 (linearized)
pMCSG70
pDB.GST
pMCSG21 (linearized)
pMCSG46
pMCSG70 (linearized)
pDB.His.MBP
pMCSG22
pMCSG46 (linearized)
pMCSG71
pDB.His.NusA
pMCSG22 (linearized)
pMCSG48
pMCSG71 (linearized)
pDB.His.TRX
pMCSG23
pMCSG48 (linearized)
pMCSG73
pDB.TRX
pMCSG23 (linearized)
pMCSG49
pMCSG73 (linearized)
pEU-GST-FV
pMCSG24
pMCSG49 (linearized)
pMCSG75
pEU-His-FV
pMCSG24 (linearized)
pMCSG50
pMCSG75 (linearized)
pEU-HSBC
pMCSG25
pMCSG50 (linearized)
pMCSG76
pFB-LIC-Bse
pMCSG25 (linearized)
pMCSG51
pMCSG76 (linearized)
pFB-LIC-Bse (linearized)
pMCSG26
pMCSG51 (linearized)
pMCSG77
pH3c-LIC
pMCSG26 (linearized)
pMCSG52
pMCSG77 (linearized)
pH3c-LIC (linearized)
pMCSG27
pMCSG52 (linearized)
pMCSG79
pHM3C-LIC
pMCSG27 (linearized)
pMCSG53
pMCSG79 (linearized)
pHM3C-LIC (linearized)
pMCSG28
pMCSG53 (linearized)
pMCSG8
pLIC-CH
pMCSG28 (linearized)
pMCSG54
pMCSG8 (linearized)
pLIC-CH (linearized)
pMCSG29
pMCSG54 (linearized)
pMCSG81
pLIC-EGFP
pMCSG29 (linearized)
pMCSG55
pMCSG81 (linearized)
pLIC-EGFP (linearized)
pMCSG30
pMCSG55 (linearized)
pMCSG9
pLIC-SGC1
pMCSG30 (linearized)
pMCSG56
pMCSG9 (linearized)
pLIC-SGC1 (linearized)
pMCSG31
pMCSG56 (linearized)
pNH-TrxT
pMCentr1
pMCSG31 (linearized)
pMCSG57
pNH-TrxT (linearized)
pMCentr1 (linearized)
pMCSG32
pMCSG57 (linearized)
pNIC-CH
pMCentr2
pMCSG32 (linearized)
pMCSG58
pNIC-CH (linearized)
pMCentr2 (linearized)
pMCSG33
pMCSG58 (linearized)
pNIC-CTHF
pMCentr3
pMCSG33 (linearized)
pMCSG59
pNIC-CTHF (linearized)
pMCentr3 (linearized)
pMCSG34
pMCSG59 (linearized)
pNIC-Zb
pMCSG10
pMCSG34 (linearized)
pMCSG60
pNIC-Zb (linearized)
pMCSG10 (linearized)
pMCSG34B
pMCSG60 (linearized)
pNIC28-Bsa4
pMCSG11
pMCSG34B (linearized)
pMCSG61
pNIC28-Bsa4 (linearized)
pMCSG11 (linearized)
pMCSG35
pMCSG61 (linearized)
pNYCOMPS-LIC-FH10T+ (N term) - linearized
pMCSG12
pMCSG35 (linearized)
pMCSG62
pNYCOMPS-LIC-FH10T+ (N term)
pMCSG12 (linearized)
pMCSG35B
pMCSG62 (linearized)
pSpeedET
pMCSG13
pMCSG35B (linearized)
pMCSG63
pVP13
pMCSG13 (linearized)
pMCSG37
pMCSG63 (linearized)
pVP16
pMCSG14
pMCSG37 (linearized)
pMCSG64
pVP33A
pMCSG14 (linearized)
pMCSG38
pMCSG64 (linearized)
pVP33K
pMCSG17
pMCSG38 (linearized)
pMCSG65
pVP56A
pMCSG17 (linearized)
pMCSG38C
pMCSG65 (linearized)
pVP56K
pMCSG18
pMCSG38C (linearized)
pMCSG66
pVP65K
pMCSG18 (linearized)
pMCSG39
pMCSG66 (linearized)
pVP68K
pMCSG19
pMCSG39 (linearized)
pMCSG67
pVP81K
pMCSG19 (linearized)
pMCSG40
pMCSG67 (linearized)
pVP91A
pMCSG19B
pMCSG40 (linearized)
pMCSG68

Plasmid Sets

Basic Cloning Vectors
I.M.A.G.E. Consortium Plasmids
pET & Duet Vectors (Novagen)
TA and GC Cloning Vectors
Coronavirus Resources
Insect Cell Vectors
pGEX Vectors (GE Healthcare)
TOPO Cloning Vectors
CRISPR Plasmids
Luciferase Vectors
Plant Vectors
Viral Expression & Packaging Vectors
Fluorescent Protein Genes & Plasmids
Lucigen Vectors
Qiagen Vectors
Yeast Plasmids
Gateway® Cloning Vectors
Mammalian Expression Vectors
Structural Genomics Vectors

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