pSpot4

Bacterial plasmid for inducible expression of fusion proteins with an N-terminal Spot-Tag®.

Sequence Author: ChromoTek

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HomePlasmidsBasic Cloning VectorspSpot4
lac operator MfeI (2900) PvuII (2813) BspQI - SapI (2750) AflIII - PciI (2633) PspFI (2333) BseYI (2329) AlwNI (2224) AhdI (1745) BsrFI (1660) BglI (1627) NmeAIII (1598) FspI (1522) T7 promoter XbaI (47) RBS BssHII (102) Spot-Tag BamHI (127) EcoRI (133) Eco53kI (141) BanII - SacI (143) SalI (146) AccI (147) HincII (148) HindIII (152) EagI - NotI (159) AvaI - BsoBI - PaeR7I - PspXI - XhoI (167) BlpI (235) StyI (257) NdeI (571) BstAPI (575) PfoI (706) EcoO109I (765) ZraI (824) AatII (826) ScaI (1264) TsoI (1347) PvuI (1376) pSpot4 2972 bp
MfeI  (2900)
1 site		 C A A T T G G T T A A C
PvuII  (2813)
1 site		 C A G C T G G T C G A C
BspQI  (2750)
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  (2750)
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.
AflIII  (2633)
1 site		 A C R Y G T T G Y R C A
Sticky ends from different AflIII sites may not be compatible.
PciI  (2633)
1 site		 A C A T G T T G T A C A
PciI is inhibited by nonionic detergents.
PspFI  (2333)
1 site		 C C C A G C G G G T C G
BseYI  (2329)
1 site		 C C C A G C G G G T C G
After cleavage, BseYI can remain bound to DNA and alter its electrophoretic mobility.
AlwNI  (2224)
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.
AhdI  (1745)
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.
BsrFI  (1660)
1 site		 R C C G G Y Y G G C C R
Cleavage may be enhanced when more than one copy of the BsrFI recognition sequence is present.
After cleavage, BsrFI can remain bound to DNA and alter its electrophoretic mobility.
BglI  (1627)
1 site		 G C C N N N N N G G C C G G N N N N N C C G
Sticky ends from different BglI sites may not be compatible.
NmeAIII  (1598)
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).
FspI  (1522)
1 site		 T G C G C A A C G C G T
XbaI  (47)
1 site		 T C T A G A A G A T C T
BssHII  (102)
1 site		 G C G C G C C G C G C G
BssHII is typically used at 50°C, but is 75% active at 37°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.
EcoRI  (133)
1 site		 G A A T T C C T T A A G
Eco53kI  (141)
1 site		 G A G C T C C T C G A G
BanII  (143)
1 site		 G R G C Y C C Y C G R G
Sticky ends from different BanII sites may not be compatible.
SacI  (143)
1 site		 G A G C T C C T C G A G
SalI  (146)
1 site		 G T C G A C C A G C T G
AccI  (147)
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.
HincII  (148)
1 site		 G T Y R A C C A R Y T G
HindIII  (152)
1 site		 A A G C T T T T C G A A
EagI  (159)
1 site		 C G G C C G G C C G G C
NotI  (159)
1 site		 G C G G C C G C C G C C G G C G
AvaI  (167)
1 site		 C Y C G R G G R G C Y C
Sticky ends from different AvaI sites may not be compatible.
BsoBI  (167)
1 site		 C Y C G R G G R G C Y C
Sticky ends from different BsoBI sites may not be compatible.
BsoBI is typically used at 37°C, but can be used at temperatures up to 65°C.
PaeR7I  (167)
1 site		 C T C G A G G A G C T C
PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (167)
1 site		 V C T C G A G B B G A G C T C V
XhoI  (167)
1 site		 C T C G A G G A G C T C
BlpI  (235)
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.
StyI  (257)
1 site		 C C W W G G G G W W C C
Sticky ends from different StyI sites may not be compatible.
NdeI  (571)
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.
BstAPI  (575)
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.
PfoI  (706)
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.
EcoO109I  (765)
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.
ZraI  (824)
1 site		 G A C G T C C T G C A G
AatII  (826)
1 site		 G A C G T C C T G C A G
ScaI  (1264)
1 site		 A G T A C T T C A T G A
TsoI  (1347)
1 site		 T A R C C A ( N ) 9 N N A T Y G G T ( N ) 9
Sticky ends from different TsoI sites may not be compatible.
After cleavage, TsoI can remain bound to DNA and alter its electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
PvuI  (1376)
1 site		 C G A T C G G C T A G C
AmpR
958 .. 1818  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   958 .. 1026  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
958 .. 1818  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   1027 .. 1818  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
958 .. 1818  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
1989 .. 2577  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
1989 .. 2577  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
AmpR promoter
853 .. 957  =  105 bp
AmpR promoter
853 .. 957  =  105 bp
MCS
127 .. 173  =  47 bp
multiple cloning site
MCS
127 .. 173  =  47 bp
multiple cloning site
ATG
88 .. 90  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
88 .. 90  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
Spot-Tag
91 .. 126  =  36 bp
12 amino acids  =  1.4 kDa
Product: engineered Spot-Tag® peptide that binds with high affinity to a nanobody
Spot-Tag
91 .. 126  =  36 bp
12 amino acids  =  1.4 kDa
Product: engineered Spot-Tag® peptide that binds with high affinity to a nanobody
lac operator
20 .. 44  =  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
20 .. 44  =  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).
RBS
59 .. 81  =  23 bp
efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)
RBS
59 .. 81  =  23 bp
efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)
T7 promoter
1 .. 19  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
1 .. 19  =  19 bp
promoter for bacteriophage T7 RNA polymerase
lac operator
2903 .. 2919  =  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
2903 .. 2919  =  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:  958 .. 1818  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  1422 .. 1688  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  2737 .. 403  =  639 bp
ORF:  212 amino acids  =  23.6 kDa
ORF:  325 .. 549  =  225 bp
ORF:  74 amino acids  =  8.5 kDa
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Individual Sequences & Maps

BlueScribe
pEASY-T1 (linearized)
pJET1.2
pSMART LCAmp
CAG promoter
pEASY-T1 Simple (linearized)
pJET1.2 blunt
pSMART LCAmp (linearized)
chicken beta-actin promoter
pEASY-T3 (linearized)
pJW168
pSMART LCKan
CLIP-tag
pEASY-T5 Zero (linearized)
pKF 18k-2
pSMART LCKan (linearized)
CMV promoter
pEX-1
pKF 19k-2
pSP64 Poly(A)
GAL1 promoter
pEX-A
pKOV
pSP72
GST
pEX-C-GST
pLATE11 (linearized)
pSP73
HaloTag
pEX-C-His
pLATE31 (linearized)
pSpark Done (linearized)
HSV TK promoter
pEX-K
pLATE51 (linearized)
pSpark I (linearized)
lac operator
pEX-N-GST
pLATE52 (linearized)
pSpark II (linearized)
lac promoter
pEX-N-His
pLEXX-AK
pSpark III (linearized)
lac UV5 promoter
pEX-N-His-GST
pLPS-B (linearized)
pSpark IV (linearized)
lacZ
pEX1
pMAL-c5E
pSpark V (linearized)
lambda
pEX2
pMAL-c5G
pSpot1
LITMUS28
pEX3
pMAL-c5X
pSpot2
LITMUS28i
pEZSeq-Amp
pMAL-c5X-His
pSpot3
LITMUS29
pEZSeq-Amp (linearized)
pMAL-c6T
pSpot4
LITMUS38
pEZSeq-Kan
pMAL-p5E
pSTBlue-1
LITMUS38i
pEZSeq-Kan (linearized)
pMAL-p5G
pSTV28
LITMUS39
pEZ BAC
pMAL-p5X
pSTV29
M13mp18
pEZ BAC (BamHI linearized)
pMCS5
pT7Blue
M13mp19
pEZ BAC (HpaI linearized)
pMiniT
pT7Blue-2
MBP
pF1A T7
pMiniT (linearized)
pT7Blue-3
pACYC177
pF1K T7
pMiniT 2.0
pT7CFE1-CFtag
pACYC184
pF25A ICE T7
pMiniT 2.0 (linearized)
pT7CFE1-CGFP-HA-His
pAN7-1
pF25K ICE T7
pMMB190
pT7CFE1-CGST-HA-His
pANT7_cGST
pF3A WG (BYDV)
pMMB206
pT7CFE1-CHA
pANT7_nHA
pF3K WG (BYDV)
pMMB207
pT7CFE1-CHis
pAUR316
pFC20A HaloTag T7 SP6
pMMB208
pT7CFE1-CMyc
pBC KS(+)
pFC20K HaloTag T7 SP6
pMMB66EH
pT7CFE1-NFtag
pBC KS(-)
pFC30A His6HaloTag T7
pMMB66HE
pT7CFE1-NHA
pBC SK(+)
pFC30K His6HaloTag T7
pMMB67EH
pT7CFE1-NHA-CHis
pBC SK(-)
pFC7A (HQ)
pMMB67HE
pT7CFE1-NHis
pBeloBAC11
pFC7K (HQ)
pNEB193
pT7CFE1-NHis-GST
pBluescript II KS(+)
pFN18A HaloTag T7
pNEB206A (linearized)
pT7CFE1-NHis-GST-CHA
pBluescript II KS(-)
pFN18K HaloTag T7
pOSIP-CH
pT7CFE1-NMyc
pBluescript II SK(+)
pFN19A HaloTag T7 SP6
pOSIP-CO
pTOP Blunt V2 (linearized)
pBluescript II SK(-)
pFN19K HaloTag T7 SP6
pOSIP-CT
pTrcHis A
pBluescript KS(+)
pFN29A His6HaloTag T7
pOSIP-KC
pTrcHis B
pBluescript KS(-)
pFN29K His6HaloTag T7
pOSIP-KH
pTrcHis C
pBluescript SK(+)
pFN2A (GST)
pOSIP-KL
pTrcHis2 A
pBluescript SK(-)
pFN2K (GST)
pOSIP-KO
pTrcHis2 B
pBR322
pFN6A (HQ)
pOSIP-KP
pTrcHis2 C
pBS KS(+)
pFN6K (HQ)
pOSIP-KT
pTV 118N
pBS KS(-)
pGEM-11Zf(+)
pOSIP-TH
pTWV228
pBS SK(+)
pGEM-11Zf(-)
pOSIP-TO
pTXB1
pBS SK(-)
pGEM-3Z
pOSIP-TT
pTYB21
pBS(+)
pGEM-3Zf(+)
pPCR-Script Amp SK(+)
pType IIs
pBS(-)
pGEM-3Zf(-)
pPCR-Script Cam SK(+)
pTZ19R
pBSK(+) Simple-Amp
pGEM-4Z
pPTR I
pTZ19U
pBSK(+) Simple-Kan
pGEM-5Zf(+)
pPTR II
pTZ57R
pcDNA3
pGEM-7Zf(+)
pRACE (linearized)
pUAST
pcDNA3.1(+)
pGEM-7Zf(-)
pRANGER-BTB-1
pUC118
pcDNA3.1(-)
pGEM-9Zf(-)
pRANGER-BTB-2
pUC119
pCold I
pGEM-T
pRANGER-BTB-3
pUC18
pCold II
pGEM-T (linearized)
pRANGER-BTB-5
pUC19
pCold III
pGEM-T Easy
pRHA-109
pUC19L
pCold IV
pGEM-T Easy (linearized)
pRHA-113
pUC57
pCold TF
pGGA
pRHA-67
pUC57-Kan
pCR-XL-2-TOPO
pGGAselect
pRSET A
pUC57-Simple
pCR-XL-2-TOPO (linearized)
PGK promoter
pRSET B
pUCmu
pDD
pGS-21a
pRSET C
pVLT31
pDNR-Dual
pH6HTC His6HaloTag T7
pSB1C3
pVLT33
pDNR-LIB
pH6HTN His6HaloTag T7
pSC101
pVLT35
pE-SUMOpro Amp
pHP45(omega)
pSEAP2-Basic
pYES1L
pE-SUMOpro Kan
pHP45(omega)-Km
pSF-Core
pZErO-2
pE-SUMOpro3 Amp
pHP45(omega)-Tc
pSMART BAC v2.0
SNAP-tag
pE-SUMOpro3 Kan
pHSG298
pSMART BAC v2.0 (BamHI linearized)
SP6 promoter
pE-SUMOstar Amp
pHSG299
pSMART BAC v2.0 (EcoRI linearized)
SV40 promoter
pE-SUMOstar Kan
pHSG396
pSMART BAC v2.0 (HindIII linearized)
T3 promoter
pEASY-Blunt (linearized)
pHSG398
pSMART FOS v2.0 (linearized)
T5 promoter
pEASY-Blunt E1 (linearized)
pHY300PLK
pSMART HCAmp
T7 promoter
pEASY-Blunt E2 (linearized)
pICOz
pSMART HCAmp (linearized)
tac promoter
pEASY-Blunt Simple (linearized)
pInvRecA
pSMART HCKan
trc promoter
pEASY-Blunt Zero (linearized)
pJAZZ-OC
pSMART HCKan (linearized)
phiX174
pEASY-Blunt3 (linearized)
pJAZZ-OK

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