LOCUS       sequence-231827-        7212 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses N-terminus of split luciferase fused to antiparallel 
            coiled-coil AP4 connected with autoinhibitory coil P3mS and TEV 
            cleavage site in antiparallel harpin linker/for SPOC logic.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7212)
  AUTHORS   Fink T, Lonzaric J, Praznik A, Plaper T, Merljak E, Leben K, Jerala 
            N, Lebar T, Strmsek Z, Lapenta F, Bencina M, Jerala R
  TITLE     Design of fast proteolysis-based signaling and logic circuits in 
            mammalian cells.
  JOURNAL   Nat Chem Biol. 2019 Feb;15(2):115-122. doi: 
            10.1038/s41589-018-0181-6. Epub 2018 Dec 10.
  PUBMED    30531965
REFERENCE   2  (bases 1 to 7212)
  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: 
            "10.1038/s41589-018-0181-6"; journalName: "Nat Chem Biol"; date: 
            "2019-02"; volume: "15"; issue: "2"; pages: "115-122"
COMMENT     Sequence Label: nLuc_AP4_TEVs_P3mS
FEATURES             Location/Qualifiers
     source          1..7212
                     /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      944..953
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             953..982
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2609..2629
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     promoter        complement(2765..2783)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2766..2783)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2803..2820)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2809..3033
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3079..3507
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3166..3185)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3376..3397
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3516..3536)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3521..3850
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3701..3836
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3763..3782
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3917..4711
                     /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(3971..3990)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4581..4600
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4885..5006
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4922..4941)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4976..4995
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5055..5071)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5055..5071)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5068..5090)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5079..5095
                     /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(5103..5109,5110..5127,5128..5133))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5148..5169
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5286..5303)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5457..6045)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5537..5556)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6216..7007,7008..7076))
                     /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     6839..6858
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7077..7181)
                     /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 gcttggtacc
      901 gagctcggat ccactagtaa cggccgccag tgtgctggaa ttcgccacca tggagcagaa
      961 gctgatcagc gaggaggacc tgggttctgg agaagatgcc aaaaacatta agaagggccc
     1021 agcgccattc tacccactcg aagacgggac cgccggcgag cagctgcaca aagccatgaa
     1081 gcgctacgcc ctggtgcccg gcaccatcgc ctttaccgac gcacatatcg aggtggacat
     1141 tacctacgcc gagtacttcg agatgagcgt tcggctggca gaagctatga agcgctatgg
     1201 gctgaataca aaccatcgga tcgtggtgtg cagcgagaat agcttgcagt tcttcatgcc
     1261 cgtgttgggt gccctgttca tcggtgtggc tgtggcccca gctaacgaca tctacaacga
     1321 gcgcgagctg ctgaacagca tgggcatcag ccagcccacc gtcgtattcg tgagcaagaa
     1381 agggctgcaa aagatcctca acgtgcaaaa gaagctaccg atcatacaaa agatcatcat
     1441 catggatagc aagaccgact accagggctt ccaaagcatg tacaccttcg tgacttccca
     1501 tttgccaccc ggcttcaacg agtacgactt cgtgcccgag agcttcgacc gggacaaaac
     1561 catcgccctg atcatgaaca gtagtggcag taccggattg cccaagggcg tagccctacc
     1621 gcaccgcacc gcttgtgtcc gattcagtca tgcccgcgac cccatcttcg gcaaccagat
     1681 catccccgac accgctatcc tcagcgtggt gccatttcac cacggcttcg gcatgttcac
     1741 cacgctgggc tacttgatct gcggctttcg ggtcgtgctc atgtaccgct tcgaggagga
     1801 gctattcttg cgcagcttgc aagactataa gattcaatct gccctgctgg tgcccacact
     1861 atttagcttc ttcgctaaga gcactctcat cgacaagtac gacctaagca acttgcacga
     1921 gatcgccagc ggcggggcgc cgctcagcaa ggaggtaggt gaggccgtgg ccaaacgctt
     1981 ccacctacca ggcatccgcc agggctacgg cctgacagaa acaaccagcg ccattctgat
     2041 cacccccgaa ggggacgaca agcctggcgc agtaggcaag gtggtgccct tcttcgaggc
     2101 taaggtggtg gacttggaca ccggtaagac actgggtgtg aaccagcgcg gcgagctgtg
     2161 cgtccgtggc cccatgatca tgagcggcta cgttaacaac cccgaggcta caaacgctct
     2221 catcgacaag gacggctggc tgcacagcgg cgacatcgcc tactgggacg aggacgagca
     2281 cttcttcatc gtggaccggc tgaagagcct gatcaaatac aagggctacc aggtagcccc
     2341 agccgaactg gagagcatcc tgctgcaaca ccccaacatc ttcgacgccg gggtcgccgg
     2401 cctgcccgac gacgatgccg gcgagctgcc cgccgcagtc gtcgtgctgg aacacggtaa
     2461 aggcggctct ggcggcggct ccggaggctc ttcacccgag gacgagcttg ctgctaatga
     2521 agaggagttg cagcaaaatg aacaaaagtt ggctcaaatt aagcaaaaac ttcaagctat
     2581 caaatacggt ggcggctctg gcggcggcga aaatttgtat ttccagagtg gcggctccgg
     2641 aggctcttcc cccgaggacg agatccagca gctcgaggag gagatatccc agcttgagca
     2701 gaagaactcc cagcttaagg agaagaacca gcagctgaag tacggctaat ctagagggcc
     2761 ctattctata gtgtcaccta aatgctagag ctcgctgatc agcctcgact gtgccttcta
     2821 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     2881 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     2941 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     3001 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
     3061 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     3121 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     3181 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     3241 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     3301 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     3361 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     3421 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     3481 tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg
     3541 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     3601 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3661 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     3721 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     3781 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     3841 cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg
     3901 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     3961 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     4021 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     4081 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     4141 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     4201 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     4261 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     4321 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     4381 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     4441 ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     4501 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     4561 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     4621 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     4681 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     4741 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     4801 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     4861 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     4921 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     4981 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     5041 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     5101 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     5161 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     5221 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     5281 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     5341 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     5401 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     5461 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5521 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     5581 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     5641 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     5701 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     5761 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5821 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5881 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5941 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     6001 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     6061 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     6121 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     6181 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     6241 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     6301 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     6361 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     6421 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     6481 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     6541 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     6601 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     6661 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     6721 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     6781 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     6841 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6901 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6961 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     7021 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     7081 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     7141 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     7201 ccacctgacg tc
//
