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Plasmid Files

pKindling-Red-B

Vector for expressing KFP-Red in bacteria.

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pKindling-Red-B.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Evrogen
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BbsI (4092) EcoO109I (4090) AatII (4036) ZraI (4034) XmnI (3713) PvuI (3484) FspI (3336) AseI (3286) NmeAIII (3262) AhdI (3114) PciI (2221) BspQI - SapI (2105) BstAPI (2045) PaeR7I - XhoI (1) PsiI (49) MfeI (59) BamHI (127) BsrGI (479) BanII (557) NgoMIV (632) NaeI (634) BsgI (637) AarI (659) HindIII (831) BlpI (843) NheI (951) BmtI (955) Bpu10I (976) PvuII (1103) BspEI (1199) PasI - PflMI * (1435) MscI (1470) XbaI (1808) PfoI (1865) PflFI - Tth111I (1968) AccI (1993) BstZ17I (1994) NdeI (2044) pKindling-Red-B 4106 bp
BbsI  (4092)
1 site
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
EcoO109I  (4090)
1 site
R G G N C C Y Y C C N G G R

Sticky ends from different EcoO109I sites may not be compatible.
AatII  (4036)
1 site
G A C G T C C T G C A G
ZraI  (4034)
1 site
G A C G T C C T G C A G
XmnI  (3713)
1 site
G A A N N N N T T C C T T N N N N A A G
PvuI  (3484)
1 site
C G A T C G G C T A G C
FspI  (3336)
1 site
T G C G C A A C G C G T
AseI  (3286)
1 site
A T T A A T T A A T T A
NmeAIII  (3262)
1 site
G C C G A G ( N ) 18-19 N N C G G C T C ( N ) 18-19

Efficient cleavage requires at least two copies of the NmeAIII recognition sequence.
Sticky ends from different NmeAIII sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
AhdI  (3114)
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.
PciI  (2221)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (2105)
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  (2105)
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.
BstAPI  (2045)
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.
PaeR7I  (1)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (1)
1 site
C T C G A G G A G C T C
PsiI  (49)
1 site
T T A T A A A A T A T T
MfeI  (59)
1 site
C A A T T G G T T A A C
BamHI  (127)
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.
BsrGI  (479)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
BanII  (557)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
NgoMIV  (632)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV recognition sequence.
NaeI  (634)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI recognition sequence.
BsgI  (637)
1 site
G T G C A G ( N ) 14 N N C A C G T C ( N ) 14

Efficient cleavage requires at least two copies of the BsgI recognition sequence.
Sticky ends from different BsgI sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
AarI  (659)
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.
HindIII  (831)
1 site
A A G C T T T T C G A A
BlpI  (843)
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.
NheI  (951)
1 site
G C T A G C C G A T C G
BmtI  (955)
1 site
G C T A G C C G A T C G
Bpu10I  (976)
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.
PvuII  (1103)
1 site
C A G C T G G T C G A C
BspEI  (1199)
1 site
T C C G G A A G G C C T
PasI  (1435)
1 site
C C C W G G G G G G W C C C

Sticky ends from different PasI sites may not be compatible.
PflMI  (1435)
1 site
C C A N N N N N T G G G G T N N N N N A C C
* Blocked by Dcm methylation.
Sticky ends from different PflMI sites may not be compatible.
MscI  (1470)
1 site
T G G C C A A C C G G T
XbaI  (1808)
1 site
T C T A G A A G A T C T
PfoI  (1865)
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.
PflFI  (1968)
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  (1968)
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.
AccI  (1993)
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.
BstZ17I  (1994)
1 site
G T A T A C C A T A T G
NdeI  (2044)
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.
AmpR
3041 .. 3901  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   3041 .. 3832  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3041 .. 3901  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   3833 .. 3901  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3041 .. 3901  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
KFP-Red
133 .. 831  =  699 bp
232 amino acids  =  25.8 kDa
Product: photoactivatable nonfluorescent protein that becomes red fluorescent upon exposure to green light
mammalian codon-optimized
KFP-Red
133 .. 831  =  699 bp
232 amino acids  =  25.8 kDa
Product: photoactivatable nonfluorescent protein that becomes red fluorescent upon exposure to green light
mammalian codon-optimized
CmR
990 .. 1649  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
990 .. 1649  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
2282 .. 2870  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2282 .. 2870  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
AmpR promoter
3902 .. 4006  =  105 bp
AmpR promoter
3902 .. 4006  =  105 bp
lambda t0 terminator
852 .. 946  =  95 bp
transcription terminator from phage lambda
lambda t0 terminator
852 .. 946  =  95 bp
transcription terminator from phage lambda
rrnB T1 terminator
1714 .. 1800  =  87 bp
transcription terminator T1 from the E. coli rrnB gene
rrnB T1 terminator
1714 .. 1800  =  87 bp
transcription terminator T1 from the E. coli rrnB gene
T5 promoter
10 .. 54  =  45 bp
   Segment 1:  
   10 .. 24  =  15 bp
bacteriophage T5 promoter for E. coli RNA polymerase, with embedded lac operator
T5 promoter
10 .. 54  =  45 bp
   Segment 2:  -35  
   25 .. 30  =  6 bp
bacteriophage T5 promoter for E. coli RNA polymerase, with embedded lac operator
T5 promoter
10 .. 54  =  45 bp
   Segment 3:  
   31 .. 47  =  17 bp
bacteriophage T5 promoter for E. coli RNA polymerase, with embedded lac operator
T5 promoter
10 .. 54  =  45 bp
   Segment 4:  -10  
   48 .. 54  =  7 bp
bacteriophage T5 promoter for E. coli RNA polymerase, with embedded lac operator
T5 promoter
10 .. 54  =  45 bp
4 segments
bacteriophage T5 promoter for E. coli RNA polymerase, with embedded lac operator
lac operator
62 .. 78  =  17 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
62 .. 78  =  17 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).
ATG
115 .. 117  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
115 .. 117  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
lac operator
30 .. 46  =  17 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
30 .. 46  =  17 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).
ORF:  115 .. 831  =  717 bp
ORF:  238 amino acids  =  26.4 kDa
ORF:  990 .. 1649  =  660 bp
ORF:  219 amino acids  =  25.7 kDa
ORF:  3171 .. 3437  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  81 .. 359  =  279 bp
ORF:  92 amino acids  =  10.4 kDa
ORF:  3041 .. 3901  =  861 bp
ORF:  286 amino acids  =  31.5 kDa
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