LOCUS       sequence-307410-        6293 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes C-terminal (substrate) fragment of FRET-based bimolecular 
            PKC activity reporter (bimCKAR); cytosol targeted. Use in 
            conjunction with pcDNA3-Cerulean-FHA1-NES.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6293)
  AUTHORS   Herbst KJ, Allen MD, Zhang J
  TITLE     Luminescent kinase activity biosensors based on a versatile 
            bimolecular switch.
  JOURNAL   J Am Chem Soc. 2011 Apr 20;133(15):5676-9. doi: 10.1021/ja1117396. 
            Epub 2011 Mar 25.
  PUBMED    21438554
REFERENCE   2  (bases 1 to 6293)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1021/ja1117396";
            journalName: "J Am Chem Soc"; date: "2011-04-20- 20"; volume: "133";
            issue: "15"; pages: "5676-9"
COMMENT     Sequence Label: pcDNA3-PKCsub-YPet-NES
FEATURES             Location/Qualifiers
     source          1..6293
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             922..939
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             943..975
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(979..987,988..1002)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     primer_bind     complement(1384..1403)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             1801..1827
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     promoter        complement(1846..1864)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1847..1864)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1884..1901)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1890..2114
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2160..2588
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2247..2266)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2457..2478
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2597..2617)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2602..2931
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2782..2917
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2844..2863
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2998..3792
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3052..3071)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3662..3681
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3966..4087
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4003..4022)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4057..4076
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4136..4152)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4136..4152)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4149..4171)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4160..4176
                     /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)."
     promoter        complement(join(4184..4190,4191..4208,4209..4214))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4229..4250
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4367..4384)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4538..5126)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4618..4637)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5297..6088,6089..6157))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5920..5939
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6158..6262)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgcggcc
      901 gccaccatgg tgcggggttc tcatcatcat catcatcatg gtatggctag catgactggt
      961 ggacagcaaa tgggtcggga tctgtacgac gatgacgata aggatcccat gcgattcaga
     1021 agattccaga cgttgaagga caaggctaag gctggcggca ccggcggcag cgagctcatg
     1081 gtgagcaaag gcgaagagct gttcaccggc gtggtgccca tcctggtgga gctggacggc
     1141 gacgtgaacg gccacaagtt cagcgtgagc ggcgagggcg agggcgacgc cacctacggc
     1201 aagctgaccc tgaagctgct gtgcaccacc ggcaagctgc ccgtgccctg gcccaccctg
     1261 gtgaccaccc tgggctacgg cgtgcagtgc ttcgcccggt accccgacca catgaagcag
     1321 cacgacttct tcaagagcgc catgcccgag ggctacgtgc aggagcggac catcttcttc
     1381 aaggacgacg gcaactacaa gacccgggcc gaggtgaagt tcgagggcga caccctggtg
     1441 aaccggatcg agctgaaggg catcgacttc aaggaggaca gcaacatcct gggccacaag
     1501 ctggagtaca actacaacag ccacaacgtg tacatcaccg ccgacaagca gaagaacggc
     1561 atcaaggcca acttcaagat ccggcacaac atcgaggacg gcggcgtgca gctggccgac
     1621 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga taaccactac
     1681 ctgagctacc agagcgccct gttcaaggac cccaacgaga agcgggacca catggtgctg
     1741 ctggagttcc tgaccgccgc cggcatcacc gagggtatga atgaattgta caaagaattc
     1801 ctgccccccc tggagcgcct gaccctgtaa tctagagggc cctattctat agtgtcacct
     1861 aaatgctaga gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     1921 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     1981 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     2041 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     2101 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
     2161 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2221 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2281 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2341 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2401 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     2461 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     2521 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     2581 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     2641 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     2701 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     2761 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     2821 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     2881 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     2941 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     3001 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3061 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     3121 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     3181 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     3241 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     3301 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     3361 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     3421 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     3481 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     3541 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     3601 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     3661 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     3721 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     3781 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     3841 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     3901 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     3961 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     4021 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     4081 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     4141 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     4201 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     4261 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4321 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     4381 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     4441 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     4501 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     4561 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     4621 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     4681 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     4741 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     4801 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     4861 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     4921 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     4981 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     5041 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     5101 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5161 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5221 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5281 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5341 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5401 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5461 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5521 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     5581 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     5641 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     5701 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     5761 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     5821 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     5881 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     5941 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6001 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6061 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6121 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6181 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6241 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
