pQE-Trisystem-6

Vector for expressing C-terminally 10xHis-tagged proteins in insect and mammalian cells using the QIAgenes system.

Sequence Author: Qiagen

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BfuAI - BspMI (2331) EcoRV (2349) TspMI - XmaI (2352) SmaI (2354) BamHI (2359) EcoRI (2365) BglII (2373) AscI (2381) PstI - SbfI (2391) BstBI (2396) HindIII (2399) BfuAI - BspMI (2414) pQE-Trisystem-6 5302 bp
BfuAI  (2331)
2 sites
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  (2331)
2 sites
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.
EcoRV  (2349)
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.
TspMI  (2352)
1 site
C C C G G G G G G C C C
XmaI  (2352)
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  (2354)
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  (2359)
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.
EcoRI  (2365)
1 site
G A A T T C C T T A A G
BglII  (2373)
1 site
A G A T C T T C T A G A
AscI  (2381)
1 site
G G C G C G C C C C G C G C G G
PstI  (2391)
1 site
C T G C A G G A C G T C
SbfI  (2391)
1 site
C C T G C A G G G G A C G T C C
BstBI  (2396)
1 site
T T C G A A A A G C T T
HindIII  (2399)
1 site
A A G C T T T T C G A A
BfuAI  (2414)
2 sites
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  (2414)
2 sites
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.
Exon 1
1727 .. 1819  =  93 bp
Exon 1
1727 .. 1819  =  93 bp
Intron
1820 .. 2081  =  262 bp
Intron
1820 .. 2081  =  262 bp
Exon 2
2082 .. 2686  =  605 bp
Exon 2
2082 .. 2686  =  605 bp
AmpR
4380 .. 5240  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   4380 .. 5171  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4380 .. 5240  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   5172 .. 5240  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4380 .. 5240  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
baculovirus recombination region (lef2/ORF603)
181 .. 1008  =  828 bp
contains ORF603 and part of lef2
baculovirus recombination region (lef2/ORF603)
181 .. 1008  =  828 bp
contains ORF603 and part of lef2
baculovirus recombination region (ORF1629)
2865 .. 3570  =  706 bp
contains part of ORF1629
baculovirus recombination region (ORF1629)
2865 .. 3570  =  706 bp
contains part of ORF1629
ori
3780 .. 4367  =  588 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3780 .. 4367  =  588 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
CMV enhancer
1138 .. 1443  =  306 bp
human cytomegalovirus (CMV) immediate early enhancer
CMV enhancer
1138 .. 1443  =  306 bp
human cytomegalovirus (CMV) immediate early enhancer
chicken β-actin promoter
1449 .. 1726  =  278 bp
chicken β-actin promoter
1449 .. 1726  =  278 bp
T7 terminator
2805 .. 2852  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 terminator
2805 .. 2852  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
ORF603
343 .. 948  =  606 bp
201 amino acids  =  23.6 kDa
Product: baculovirus ORF603 protein
ORF603
343 .. 948  =  606 bp
201 amino acids  =  23.6 kDa
Product: baculovirus ORF603 protein
ORF1629
2982 .. 3570  =  589 bp
195 amino acids  =  22.3 kDa
Product: baculovirus capsid-associated protein
required for viral replication
ORF1629
2982 .. 3570  =  589 bp
195 amino acids  =  22.3 kDa
Product: baculovirus capsid-associated protein
required for viral replication
lef2
181 .. 305  =  125 bp
40 amino acids  =  4.5 kDa
Product: baculovirus late expression factor 2
lef2
181 .. 305  =  125 bp
40 amino acids  =  4.5 kDa
Product: baculovirus late expression factor 2
rabbit globin terminator
2684 .. 2795  =  112 bp
rabbit globin terminator
2684 .. 2795  =  112 bp
p10 promoter
2209 .. 2318  =  110 bp
baculovirus promoter for expression in insect cells
p10 promoter
2209 .. 2318  =  110 bp
baculovirus promoter for expression in insect cells
MCS
2347 .. 2404  =  58 bp
multiple cloning site
MCS
2347 .. 2404  =  58 bp
multiple cloning site
10xHis
2415 .. 2444  =  30 bp
10 amino acids  =  1.4 kDa
Product: 10xHis affinity tag
10xHis
2415 .. 2444  =  30 bp
10 amino acids  =  1.4 kDa
Product: 10xHis affinity tag
lac operator
2169 .. 2193  =  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
2169 .. 2193  =  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).
8xHis
2484 .. 2507  =  24 bp
8 amino acids  =  1.1 kDa
Product: 8xHis affinity tag
8xHis
2484 .. 2507  =  24 bp
8 amino acids  =  1.1 kDa
Product: 8xHis affinity tag
T7 promoter
2150 .. 2168  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
2150 .. 2168  =  19 bp
promoter for bacteriophage T7 RNA polymerase
siRNA target site
2459 .. 2477  =  19 bp
siRNA target site
2459 .. 2477  =  19 bp
stop codons
2508 .. 2518  =  11 bp
stop codons in all three reading frames
stop codons
2508 .. 2518  =  11 bp
stop codons in all three reading frames
RBS
2321 .. 2326  =  6 bp
ribosome binding site
RBS
2321 .. 2326  =  6 bp
ribosome binding site
stop codons
2445 .. 2450  =  6 bp
two tandem stop codons
stop codons
2445 .. 2450  =  6 bp
two tandem stop codons
poly(A) signal
2662 .. 2667  =  6 bp
poly(A) signal
2662 .. 2667  =  6 bp
ATG
2338 .. 2340  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
2338 .. 2340  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ORF:  1315 .. 1704  =  390 bp
ORF:  129 amino acids  =  13.9 kDa
ORF:  4510 .. 4776  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  5290 .. 305  =  318 bp
ORF:  105 amino acids  =  12.1 kDa
ORF:  1547 .. 1798  =  252 bp
ORF:  83 amino acids  =  8.7 kDa
ORF:  343 .. 948  =  606 bp
ORF:  201 amino acids  =  23.6 kDa
ORF:  387 .. 617  =  231 bp
ORF:  76 amino acids  =  9.2 kDa
ORF:  1671 .. 2060  =  390 bp
ORF:  129 amino acids  =  13.3 kDa
ORF:  2403 .. 2648  =  246 bp
ORF:  81 amino acids  =  9.0 kDa
ORF:  2982 .. 3599  =  618 bp
ORF:  205 amino acids  =  23.4 kDa
ORF:  4380 .. 5240  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
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