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pQCXIX

Self-inactivating bicistronic retroviral vector for simultaneous expression of two genes.

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pQCXIX Sequence and MappQCXIX.dna
Map and Sequence File   
Sequence Author:  Clontech
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 SspI (5964) ScaI (5640) PvuI (5530) FspI (5382) AlwNI (4683) BspQI - SapI (4151) SfiI (3973) AccI (3139) SalI (3138) BmtI (3049) CMV enhancer AscI (542) PshAI (676) AfeI (1088) BstEII (1252) Bpu10I (1455) BglII (1576) BspDI * - ClaI * (1588) XbaI (1595) CMV enhancer SbfI (2230) NotI (2243) AgeI (2250) BsiWI (2262) PacI (2273) BamHI (2278) BspEI * (2281) EcoRI (2285) PspOMI (2421) ApaI (2425) PmlI (2624) AarI (2647) DraIII (2671) PflMI (2761) BmgBI (2851) BclI * (2885) MluI (2890) PaeR7I - XhoI (2896) EcoRV (2903) PvuII (2946) NheI (3045) pQCXIX 6309 bp
SspI  (5964)
1 site
A A T A T T T T A T A A
ScaI  (5640)
1 site
A G T A C T T C A T G A
PvuI  (5530)
1 site
C G A T C G G C T A G C
FspI  (5382)
1 site
T G C G C A A C G C G T
AlwNI  (4683)
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.
BspQI  (4151)
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  (4151)
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.
SfiI  (3973)
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.
AccI  (3139)
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.
SalI  (3138)
1 site
G T C G A C C A G C T G
BmtI  (3049)
1 site
G C T A G C C G A T C G
AscI  (542)
1 site
G G C G C G C C C C G C G C G G
PshAI  (676)
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.
AfeI  (1088)
1 site
A G C G C T T C G C G A
BstEII  (1252)
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.
Bpu10I  (1455)
1 site
C C T N A G C G G A N T C G

Efficient cleavage requires at least two copies of the Bpu10I
recognition sequence.
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.
BglII  (1576)
1 site
A G A T C T T C T A G A
BspDI  (1588)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (1588)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
XbaI  (1595)
1 site
T C T A G A A G A T C T
SbfI  (2230)
1 site
C C T G C A G G G G A C G T C C
NotI  (2243)
1 site
G C G G C C G C C G C C G G C G
AgeI  (2250)
1 site
A C C G G T T G G C C A

AgeI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
BsiWI  (2262)
1 site
C G T A C G G C A T G C

BsiWI is typically used at 55°C, but is 50% active at 37°C.
PacI  (2273)
1 site
T T A A T T A A A A T T A A T T
BamHI  (2278)
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.
BspEI  (2281)
1 site
T C C G G A A G G C C T
* Blocked by Dam methylation.
EcoRI  (2285)
1 site
G A A T T C C T T A A G
PspOMI  (2421)
1 site
G G G C C C C C C G G G
ApaI  (2425)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PmlI  (2624)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
AarI  (2647)
1 site
C A C C T G C ( N ) 4 G T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the AarI
recognition sequence.
Sticky ends from different AarI sites may not be compatible.
After cleavage, AarI can remain bound to DNA and alter its
electrophoretic mobility.
DraIII  (2671)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
PflMI  (2761)
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.
BmgBI  (2851)
1 site
C A C G T C G T G C A G

This recognition sequence is asymmetric, so ligating blunt ends
generated by BmgBI will not always regenerate a BmgBI site.
BclI  (2885)
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.
MluI  (2890)
1 site
A C G C G T T G C G C A
PaeR7I  (2896)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (2896)
1 site
C T C G A G G A G C T C
EcoRV  (2903)
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.
PvuII  (2946)
1 site
C A G C T G G T C G A C
NheI  (3045)
1 site
G C T A G C C G A T C G
AmpR
5087 .. 5947  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   5087 .. 5878  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
5087 .. 5947  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   5879 .. 5947  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
5087 .. 5947  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
4328 .. 4916  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
4328 .. 4916  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
IRES
2308 .. 2881  =  574 bp
internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)
IRES
2308 .. 2881  =  574 bp
internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)
3' LTR (ΔU3)
3015 .. 3440  =  426 bp
self-inactivating 3' long terminal repeat (LTR) from
Moloney murine leukemia virus
3' LTR (ΔU3)
3015 .. 3440  =  426 bp
self-inactivating 3' long terminal repeat (LTR) from
Moloney murine leukemia virus
gag (truncated)
1151 .. 1567  =  417 bp
truncated MMLV gag gene lacking the start codon
gag (truncated)
1151 .. 1567  =  417 bp
truncated MMLV gag gene lacking the start codon
CMV enhancer
6238 .. 308  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
6238 .. 308  =  380 bp
human cytomegalovirus immediate early enhancer
SV40 promoter
3706 .. 4035  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
3706 .. 4035  =  330 bp
SV40 enhancer and early promoter
CMV enhancer
1604 .. 1907  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
1604 .. 1907  =  304 bp
human cytomegalovirus immediate early enhancer
CMV promoter
309 .. 512  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
309 .. 512  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
1908 .. 2111  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
1908 .. 2111  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
MMLV Ψ
751 .. 950  =  200 bp
packaging signal of Moloney murine leukemia virus
(MMLV)
MMLV Ψ
751 .. 950  =  200 bp
packaging signal of Moloney murine leukemia virus
(MMLV)
5' LTR (truncated)
513 .. 688  =  176 bp
truncated long terminal repeat from Moloney murine
sarcoma virus
5' LTR (truncated)
513 .. 688  =  176 bp
truncated long terminal repeat from Moloney murine
sarcoma virus
AmpR promoter
5948 .. 6052  =  105 bp
AmpR promoter
5948 .. 6052  =  105 bp
MCS 1
2225 .. 2290  =  66 bp
multiple cloning site
MCS 1
2225 .. 2290  =  66 bp
multiple cloning site
MCS 2
2885 .. 2906  =  22 bp
multiple cloning site
MCS 2
2885 .. 2906  =  22 bp
multiple cloning site
SV40 ori
3886 .. 4021  =  136 bp
SV40 origin of replication
SV40 ori
3886 .. 4021  =  136 bp
SV40 origin of replication
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