pGFP-B-RS

HuSH-29 retroviral vector with kanamycin and blasticidin resistance markers and a TurboGFP gene, for expressing an shRNA from the U6 promoter.

Sequence Author: OriGene

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EcoRI (1) CsiI - SexAI * (7189) BstEII (7061) Bsu36I (6763) PfoI (6516) HpaI (5031) MfeI (5023) BsrDI (4930) CMV enhancer SnaBI (4500) BglII (4192) XmnI (3947) ApaI (3600) DraIII (40) BamHI (334) HindIII (380) SfiI (597) StuI (643) AvrII (644) BclI * (654) BsiWI (667) BspDI - ClaI (1109) PciI (2057) ApaLI (2371) AgeI (2819) BstXI (2938) AleI (3311) SacII (3399) PmeI (3411) PaeR7I - PspXI - XhoI (3416) MluI (3434) RsrII (3441) AclI (3466) AsiSI (3475) PspOMI (3596) pGFP-B-RS 7388 bp
EcoRI  (1)
1 site
G A A T T C C T T A A G
CsiI  (7189)
1 site
A C C W G G T T G G W C C A

Sticky ends from different CsiI sites may not be compatible.
SexAI  (7189)
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.
BstEII  (7061)
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.
Bsu36I  (6763)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
PfoI  (6516)
1 site
T C C N G G A A G G N C C T

Sticky ends from different PfoI sites may not be compatible.
HpaI  (5031)
1 site
G T T A A C C A A T T G
MfeI  (5023)
1 site
C A A T T G G T T A A C
BsrDI  (4930)
1 site
G C A A T G N N C G T T A C

Sticky ends from different BsrDI sites may not be compatible.
SnaBI  (4500)
1 site
T A C G T A A T G C A T
BglII  (4192)
1 site
A G A T C T T C T A G A
XmnI  (3947)
1 site
G A A N N N N T T C C T T N N N N A A G
ApaI  (3600)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
DraIII  (40)
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.
BamHI  (334)
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.
HindIII  (380)
1 site
A A G C T T T T C G A A
SfiI  (597)
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.
StuI  (643)
1 site
A G G C C T T C C G G A
AvrII  (644)
1 site
C C T A G G G G A T C C
BclI  (654)
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.
BsiWI  (667)
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.
BspDI  (1109)
1 site
A T C G A T T A G C T A
ClaI  (1109)
1 site
A T C G A T T A G C T A
PciI  (2057)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
ApaLI  (2371)
1 site
G T G C A C C A C G T G
AgeI  (2819)
1 site
A C C G G T T G G C C A
BstXI  (2938)
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.
AleI  (3311)
1 site
C A C N N N N G T G G T G N N N N C A C
SacII  (3399)
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.
PmeI  (3411)
1 site
G T T T A A A C C A A A T T T G
PaeR7I  (3416)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (3416)
1 site
V C T C G A G B B G A G C T C V
XhoI  (3416)
1 site
C T C G A G G A G C T C
MluI  (3434)
1 site
A C G C G T T G C G C A
RsrII  (3441)
1 site
C G G W C C G G C C W G G C

Efficient cleavage requires at least two copies of the RsrII recognition sequence.
Sticky ends from different RsrII sites may not be compatible.
For full activity, add fresh DTT.
AclI  (3466)
1 site
A A C G T T T T G C A A
AsiSI  (3475)
1 site
G C G A T C G C C G C T A G C G
PspOMI  (3596)
1 site
G G G C C C C C C G G G
KanR
5091 .. 5900  =  810 bp
269 amino acids  =  30.8 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418 (Geneticin®) in eukaryotes
KanR
5091 .. 5900  =  810 bp
269 amino acids  =  30.8 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418 (Geneticin®) in eukaryotes
TurboGFP
3490 .. 4185  =  696 bp
232 amino acids  =  25.7 kDa
Product: improved green fluorescent protein from Pontellina plumata (Evdokimov et al., 2006)
mammalian codon-optimized
TurboGFP
3490 .. 4185  =  696 bp
232 amino acids  =  25.7 kDa
Product: improved green fluorescent protein from Pontellina plumata (Evdokimov et al., 2006)
mammalian codon-optimized
3' LTR
1153 .. 1746  =  594 bp
long terminal repeat from Moloney murine leukemia virus
3' LTR
1153 .. 1746  =  594 bp
long terminal repeat from Moloney murine leukemia virus
ori
2118 .. 2706  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2118 .. 2706  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
hGH poly(A) signal
2893 .. 3369  =  477 bp
human growth hormone polyadenylation signal
hGH poly(A) signal
2893 .. 3369  =  477 bp
human growth hormone polyadenylation signal
5' LTR
6034 .. 6503  =  470 bp
long terminal repeat from Moloney murine leukemia virus
5' LTR
6034 .. 6503  =  470 bp
long terminal repeat from Moloney murine leukemia virus
gag (truncated)
6960 .. 7376  =  417 bp
truncated MMLV gag gene lacking the start codon
gag (truncated)
6960 .. 7376  =  417 bp
truncated MMLV gag gene lacking the start codon
BSD
676 .. 1074  =  399 bp
132 amino acids  =  13.7 kDa
Product: blasticidin S deaminase
confers resistance to blasticidin
BSD
676 .. 1074  =  399 bp
132 amino acids  =  13.7 kDa
Product: blasticidin S deaminase
confers resistance to blasticidin
CMV enhancer
4477 .. 4856  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
4477 .. 4856  =  380 bp
human cytomegalovirus immediate early enhancer
U6 promoter
90 .. 330  =  241 bp
RNA polymerase III promoter for human U6 snRNA
U6 promoter
90 .. 330  =  241 bp
RNA polymerase III promoter for human U6 snRNA
CMV promoter
4273 .. 4476  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
4273 .. 4476  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
MMLV Ψ
6568 .. 6767  =  200 bp
packaging signal of Moloney murine leukemia virus (MMLV)
MMLV Ψ
6568 .. 6767  =  200 bp
packaging signal of Moloney murine leukemia virus (MMLV)
SV40 ori
510 .. 645  =  136 bp
SV40 origin of replication
SV40 ori
510 .. 645  =  136 bp
SV40 origin of replication
shRNA site
334 .. 385  =  52 bp
shRNA insertion site
shRNA site
334 .. 385  =  52 bp
shRNA insertion site
T7 promoter
4229 .. 4247  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
4229 .. 4247  =  19 bp
promoter for bacteriophage T7 RNA polymerase
ORF:  676 .. 1074  =  399 bp
ORF:  132 amino acids  =  13.7 kDa
ORF:  3269 .. 3565  =  297 bp
ORF:  98 amino acids  =  10.5 kDa
ORF:  3447 .. 3713  =  267 bp
ORF:  88 amino acids  =  9.3 kDa
ORF:  5091 .. 5900  =  810 bp
ORF:  269 amino acids  =  30.8 kDa
ORF:  571 .. 978  =  408 bp
ORF:  135 amino acids  =  14.7 kDa
ORF:  3478 .. 4317  =  840 bp
ORF:  279 amino acids  =  31.3 kDa
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