LOCUS       sequence-231805-        5766 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses C-terminus of split luciferase connected to P9 coil via 
            TEVs cleavable linker/for split protease-cleavable orthogonal 
            Coiled-coil (SPOC) logic.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5766)
  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 5766)
  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: P9_TEVs_cLuc
FEATURES             Location/Qualifiers
     source          1..5766
                     /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"
     CDS             1058..1078
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1274..1300
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        complement(1319..1337)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1320..1337)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1357..1374)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1363..1587
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1633..2061
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1720..1739)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1930..1951
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2070..2090)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2075..2404
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2255..2390
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2317..2336
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2471..3265
                     /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(2525..2544)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3135..3154
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3439..3560
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3476..3495)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3530..3549
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3609..3625)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3609..3625)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3622..3644)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3633..3649
                     /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(3657..3663,3664..3681,3682..3687))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3702..3723
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3840..3857)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4011..4599)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4091..4110)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4770..5561,5562..5630))
                     /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     5393..5412
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5631..5735)
                     /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 tgtctcccga
      961 agacgaaaac caagccctcg aacagaagaa cgctcaactt aaacaggaga tcgcggcact
     1021 tgaacaagag atcgcgcaac tcgaatatgg cggctctgaa aatttgtatt tccagtccgg
     1081 aaccatgacc gagaaggaga tcgtggacta tgtggccagc caggttacaa ccgccaagaa
     1141 gctgcgcggt ggtgttgtgt tcgtggacga ggtgcctaaa ggactgaccg gcaagttgga
     1201 cgcccgcaag atccgcgaga ttctcattaa ggccaagaag ggcggcaaga tcgccgtgaa
     1261 tagtggttct ggatacccat acgatgttcc agattacgct taatctagag ggccctattc
     1321 tatagtgtca cctaaatgct agagctcgct gatcagcctc gactgtgcct tctagttgcc
     1381 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     1441 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     1501 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     1561 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     1621 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     1681 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     1741 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     1801 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     1861 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     1921 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     1981 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     2041 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     2101 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     2161 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     2221 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     2281 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     2341 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     2401 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcaagaga caggatgagg
     2461 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     2521 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     2581 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     2641 gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     2701 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     2761 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     2821 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     2881 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     2941 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg
     3001 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     3061 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     3121 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     3181 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     3241 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     3301 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     3361 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     3421 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     3481 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     3541 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     3601 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     3661 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     3721 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     3781 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     3841 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3901 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3961 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4021 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4081 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4141 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4201 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4261 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4321 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4381 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     4441 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     4501 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     4561 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4621 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4681 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4741 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4801 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4861 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4921 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4981 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5041 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5101 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5161 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5221 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5281 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5341 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     5401 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5461 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5521 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5581 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5641 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5701 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5761 gacgtc
//
