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pRFP-CB-shLenti

HuSH-29 lentiviral vector with chloramphenicol and blasticidin resistance markers and a TurboRFP gene, for expressing an shRNA from the U6 promoter.

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pRFP-CB-shLenti Sequence and MappRFP-CB-shLenti.dna
Map and Sequence File   
Sequence Author:  OriGene
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 AclI (8055) PfoI (5948) PmeI (5538) PshAI (5466) BfuAI - BspMI (5353) PaeR7I - XhoI (4833) NotI (4827) MluI (4818) RsrII (4812) AscI (4781) MreI - SgrAI (4777) AsiSI (4775) CMV enhancer SgrDI (1) CMV enhancer KasI (1017) NarI (1018) SfoI (1019) PluTI (1021) XmnI (1224) BspQI - SapI (1482) EcoNI (1550) AleI (1957) KflI (2314) PspOMI (2605) ApaI (2609) EcoRI (2618) PmlI (2657) BamHI (2951) SphI (3087) NsiI (3089) StuI (3266) AvrII (3267) BsaBI * (3597) BstBI (3718) NheI (3806) BmtI (3810) XbaI (4073) pRFP-CB-shLenti 8491 bp
AclI  (8055)
1 site
A A C G T T T T G C A A
PfoI  (5948)
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.
PmeI  (5538)
1 site
G T T T A A A C C A A A T T T G
PshAI  (5466)
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.
BfuAI  (5353)
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  (5353)
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.
PaeR7I  (4833)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (4833)
1 site
C T C G A G G A G C T C
NotI  (4827)
1 site
G C G G C C G C C G C C G G C G
MluI  (4818)
1 site
A C G C G T T G C G C A
RsrII  (4812)
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.
AscI  (4781)
1 site
G G C G C G C C C C G C G C G G
MreI  (4777)
1 site
C G C C G G C G G C G G C C G C
SgrAI  (4777)
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.
AsiSI  (4775)
1 site
G C G A T C G C C G C T A G C G
SgrDI  (1)
1 site
C G T C G A C G G C A G C T G C
KasI  (1017)
1 site
G G C G C C C C G C G G
NarI  (1018)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the NarI
recognition sequence.
SfoI  (1019)
1 site
G G C G C C C C G C G G
PluTI  (1021)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI
recognition sequence.
XmnI  (1224)
1 site
G A A N N N N T T C C T T N N N N A A G
BspQI  (1482)
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  (1482)
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.
EcoNI  (1550)
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.
AleI  (1957)
1 site
C A C N N N N G T G G T G N N N N C A C
KflI  (2314)
1 site
G G G W C C C C C C W G G G

Sticky ends from different KflI sites may not be compatible.
PspOMI  (2605)
1 site
G G G C C C C C C G G G
ApaI  (2609)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
EcoRI  (2618)
1 site
G A A T T C C T T A A G
PmlI  (2657)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
BamHI  (2951)
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.
SphI  (3087)
1 site
G C A T G C C G T A C G
NsiI  (3089)
1 site
A T G C A T T A C G T A
StuI  (3266)
1 site
A G G C C T T C C G G A
AvrII  (3267)
1 site
C C T A G G G G A T C C
BsaBI  (3597)
1 site
G A T N N N N A T C C T A N N N N T A G
* Blocked by Dam methylation.
BstBI  (3718)
1 site
T T C G A A A A G C T T
NheI  (3806)
1 site
G C T A G C C G A T C G
BmtI  (3810)
1 site
G C T A G C C G A T C G
XbaI  (4073)
1 site
T C T A G A A G A T C T
TurboRFP
4839 .. 5534  =  696 bp
231 amino acids  =  26.1 kDa
Product: red fluorescent protein from Entacmaea
quadricolor

mammalian codon-optimized
TurboRFP
4839 .. 5534  =  696 bp
231 amino acids  =  26.1 kDa
Product: red fluorescent protein from Entacmaea
quadricolor

mammalian codon-optimized
CmR
7699 .. 8358  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
7699 .. 8358  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
WPRE
5675 .. 6263  =  589 bp
woodchuck hepatitis virus posttranscriptional
regulatory element
WPRE
5675 .. 6263  =  589 bp
woodchuck hepatitis virus posttranscriptional
regulatory element
ori
7034 .. 7622  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
7034 .. 7622  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
BSD
3299 .. 3697  =  399 bp
132 amino acids  =  13.7 kDa
Product: blasticidin S deaminase
confers resistance to blasticidin
BSD
3299 .. 3697  =  399 bp
132 amino acids  =  13.7 kDa
Product: blasticidin S deaminase
confers resistance to blasticidin
CMV enhancer
237 .. 616  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
237 .. 616  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
4104 .. 4483  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
4104 .. 4483  =  380 bp
human cytomegalovirus immediate early enhancer
U6 promoter
2707 .. 2947  =  241 bp
RNA polymerase III promoter for human U6 snRNA
U6 promoter
2707 .. 2947  =  241 bp
RNA polymerase III promoter for human U6 snRNA
RRE
1683 .. 1916  =  234 bp
The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to
the cytoplasm.
RRE
1683 .. 1916  =  234 bp
The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to
the cytoplasm.
CMV promoter
4484 .. 4687  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
4484 .. 4687  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
618 .. 816  =  199 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
618 .. 816  =  199 bp
human cytomegalovirus (CMV) immediate early
promoter
5' LTR (truncated)
834 .. 1014  =  181 bp
truncated 5' long terminal repeat (LTR) from HIV-1
5' LTR (truncated)
834 .. 1014  =  181 bp
truncated 5' long terminal repeat (LTR) from HIV-1
3' LTR (truncated)
6792 .. 6972  =  181 bp
truncated 3' long terminal repeat (LTR) from HIV-1
3' LTR (truncated)
6792 .. 6972  =  181 bp
truncated 3' long terminal repeat (LTR) from HIV-1
SV40 ori
3133 .. 3268  =  136 bp
SV40 origin of replication
SV40 ori
3133 .. 3268  =  136 bp
SV40 origin of replication
HIV-1 Ψ
1061 .. 1186  =  126 bp
packaging signal of human immunodeficiency virus
type 1
HIV-1 Ψ
1061 .. 1186  =  126 bp
packaging signal of human immunodeficiency virus
type 1
cPPT/CTS
2443 .. 2560  =  118 bp
central polypurine tract and central termination
sequence of HIV-1
cPPT/CTS
2443 .. 2560  =  118 bp
central polypurine tract and central termination
sequence of HIV-1
AmpR promoter
8359 .. 8463  =  105 bp
AmpR promoter
8359 .. 8463  =  105 bp
shRNA site
2951 .. 3008  =  58 bp
shRNA insertion site
shRNA site
2951 .. 3008  =  58 bp
shRNA insertion site
loxP
5586 .. 5619  =  34 bp
Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT).
loxP
5586 .. 5619  =  34 bp
Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT).
T7 promoter
4713 .. 4731  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
4713 .. 4731  =  19 bp
promoter for bacteriophage T7 RNA polymerase
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