cytoplasmic EKAR (Cerulean-Venus) vector (V007110)

Basic Vector Information

      • Vector Name:
      • cytoplasmic EKAR (Cerulean-Venus)
      • Antibiotic Resistance:
      • Ampicillin
      • Length:
      • 6758 bp
      • Type:
      • Mammalian Expression
      • Copy Number:
      • High Copy
      • Cloning Method:
      • Restriction Enzyme
      • 5' Primer:
      • CMV Forward

cytoplasmic EKAR (Cerulean-Venus) vector Vector Map

cytoplasmic EKAR (Cerulean-Venus)6758 bp3006009001200150018002100240027003000330036003900420045004800510054005700600063006600CMV enhancerCMV promoterSP6 promoterpMT2-FKozak sequencemVenusKozak sequenceSV40 poly(A) signalSV40 promoterM13 Forwardf1 oripRS-markerpGEX 3'pBRforEcoAmpR promoterAmpRoriL4440lac promoterlac operatorM13 rev

Plasmid Resuspension Protocol:

1. Centrifuge at 5,000×g for 5 min.

2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.

3. Close the tube and incubate for 10 minutes at room temperature.

4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.

5.Store the plasmid at -20 ℃.

cytoplasmic EKAR (Cerulean-Venus) vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       Exported                6758 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    cytoplasmic EKAR (Cerulean-Venus)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6758)
  AUTHORS   Harvey CD, Ehrhardt AG, Cellurale C, Zhong H, Yasuda R, Davis RJ, 
            Svoboda K.
  TITLE     A genetically encoded fluorescent sensor of ERK activity
  JOURNAL   PNAS 2008 Dec 9;105(49):19264-9
  PUBMED    19033456
REFERENCE   2  (bases 1 to 6758)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PNAS"; 
            date: "2008-12-9"; volume: "105(49)"; pages: "19264-"
FEATURES             Location/Qualifiers
     source          1..6758
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        827..845
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     827..844
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     858..877
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      933..942
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             939..1655
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2220..2936
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      3007..3016
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     polyA_signal    3020..3154
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3057..3076)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3111..3130
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        3223..3580
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(3246..3266)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3431..3566
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3493..3512
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(3600..3617)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3600..3616)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3609..3631)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3829..4284
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3916..3935)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4126..4147
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4301..4320)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4455..4477
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4530..4548)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4616..4720
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4721..5581
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRDDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4939..4958)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5752..6340
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6241..6260
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6494..6511
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        6664..6694
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6702..6718
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     6707..6729
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6726..6742
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6726..6742
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"

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