LOCUS       sequence-231830-        7715 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Circularly permutated firefly luciferase with SbMV cleavage site 
            (Luciferase reporter for detection of proteolytic activity).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7715)
  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 7715)
  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: cycLuc_SbMVs
FEATURES             Location/Qualifiers
     source          1..7715
                     /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             922..939
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3073..3120
                     /codon_start=1
                     /product="non-ORF yeast peptide conferring 
                     ubiquitin-dependent degradation"
                     /label=hCL1
                     /note="human codon-optimized"
                     /translation="ACKNWFSSLSHFVIHL"
     CDS             3124..3243
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     promoter        complement(3268..3286)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3269..3286)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3306..3323)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3312..3536
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3582..4010
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3669..3688)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3879..3900
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4019..4039)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4024..4353
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4204..4339
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4266..4285
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4420..5214
                     /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(4474..4493)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5084..5103
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5388..5509
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5425..5444)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5479..5498
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5558..5574)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5558..5574)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5571..5593)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5582..5598
                     /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(5606..5612,5613..5630,5631..5636))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5651..5672
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5789..5806)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5960..6548)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6040..6059)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6719..7510,7511..7579))
                     /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     7342..7361
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7580..7684)
                     /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 gagctcgcta gcgccaccat gcaccaccac caccaccaca tgatcaaaat agccacacgt
      961 aaatatttag gcaaacaaaa tgtctatgac attggagttg agcgcgacca taattttgca
     1021 ctcaaaaatg gcttcatagc ttctaattgt ttcaatgata cttatagata tattgacacc
     1081 gctatcctca gcgtggtgcc atttcaccac ggcttcggca tgttcaccac gctgggctac
     1141 ttgatctgcg gctttcgggt cgtgctcatg taccgcttcg aggaggagct attcttgcgc
     1201 agcttgcaag actataagat tcaatctgcc ctgctggtgc ccacactatt tagcttcttc
     1261 gctaagagca ctctcatcga caagtacgac ctaagcaact tgcacgagat cgccagcggc
     1321 ggggcgccgc tcagcaagga ggtaggtgag gccgtggcca aacgcttcca cctaccaggc
     1381 atccgccagg gctacggcct gacagaaaca accagcgcca ttctgatcac ccccgaaggg
     1441 gacgacaagc ctggcgcagt aggcaaggtg gtgcccttct tcgaggctaa ggtggtggac
     1501 ttggacaccg gtaagacact gggtgtgaac cagcgcggcg agctgtgcgt ccgtggcccc
     1561 atgatcatga gcggctacgt taacaacccc gaggctacaa acgctctcat cgacaaggac
     1621 ggctggctgc acagcggcga catcgcctac tgggacgagg acgagcactt cttcatcgtg
     1681 gaccggctga agagcctgat caaatacaag ggctaccagg tagccccagc cgaactggag
     1741 agcatcctgc tgcaacaccc caacatcttc gacgccgggg tcgccggcct gcccgacgac
     1801 gatgccggcg agctgcccgc cgcagtcgtc gtgctggaac acggtaaaac catgaccgag
     1861 aaggagatcg tggactatgt ggccagccag gttacaaccg ccaagaagct gcgcggtggt
     1921 gttgtgttcg tggacgaggt gcctaaagga ctgaccggca agttggacgc ccgcaagatc
     1981 cgcgagattc tcattaaggc caagaaggga tccgagtccg tcagcctgca aagcggctca
     2041 ggtgccaaaa acattaagaa gggcccagcg ccattctacc cactcgaaga cgggaccgcc
     2101 ggcgagcagc tgcacaaagc catgaagcgc tacgccctgg tgcccggcac catcgccttt
     2161 accgacgcac atatcgaggt ggacattacc tacgccgagt acttcgagat gagcgttcgg
     2221 ctggcagaag ctatgaagcg ctatgggctg aatacaaacc atcggatcgt ggtgtgcagc
     2281 gagaatagct tgcagttctt catgcccgtg ttgggtgccc tgttcatcgg tgtggctgtg
     2341 gccccagcta acgacatcta caacgagcgc gagctgctga acagcatggg catcagccag
     2401 cccaccgtcg tattcgtgag caagaaaggg ctgcaaaaga tcctcaacgt gcaaaagaag
     2461 ctaccgatca tacaaaagat catcatcatg gatagcaaga ccgactacca gggcttccaa
     2521 agcatgtaca ccttcgtgac ttcccatttg ccacccggct tcaacgagta cgacttcgtg
     2581 cccgagagct tcgaccggga caaaaccatc gccctgatca tgaacagtag tggcagtacc
     2641 ggattgccca agggcgtagc cctaccgcac cgcaccgctt gtgtccgatt cagtcatgcc
     2701 cgcgacccca tcttcggcaa ccagatcatc cccgccgagt actgtttaag ctatgaaacg
     2761 gaaatattga cagtagaata tggattatta ccgattggta aaattgtaga aaagcgcatc
     2821 gaatgtactg tttatagcgt tgataataat ggaaatattt atacacaacc tgtagcacaa
     2881 tggcacgatc gcggagaaca agaggtgttt gagtattgtt tggaagatgg ttcattgatt
     2941 cgggcaacaa aagaccataa gtttatgact gttgatggtc aaatgttgcc aattgatgaa
     3001 atatttgaac gtgaattgga tttgatgcgg gttgataatt tgccgaacgg cggcaagatc
     3061 gccgtcaatt ctgcttgcaa gaactggttc agtagcttaa gccactttgt gatccacctt
     3121 aacagccacg gcttccctcc cgaggtggag gagcaggccg ccggcaccct gcccatgagc
     3181 tgcgcccagg agagcggcat ggatagacac cctgctgctt gcgccagcgc caggatcaac
     3241 gtctaactgc agtctagagg gccctattct atagtgtcac ctaaatgcta gagctcgctg
     3301 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     3361 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     3421 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     3481 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     3541 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     3601 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     3661 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     3721 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     3781 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     3841 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     3901 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     3961 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     4021 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     4081 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     4141 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     4201 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     4261 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     4321 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     4381 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     4441 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     4501 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     4561 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     4621 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     4681 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     4741 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     4801 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     4861 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     4921 gccgaactgt tcgctaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     4981 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     5041 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     5101 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     5161 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     5221 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     5281 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     5341 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     5401 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     5461 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     5521 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     5581 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     5641 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5701 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5761 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5821 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5881 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5941 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     6001 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     6061 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     6121 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6181 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6241 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6301 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6361 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6421 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6481 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6541 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6601 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6661 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6721 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6781 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6841 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6901 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6961 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     7021 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     7081 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     7141 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     7201 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     7261 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     7321 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     7381 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     7441 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7501 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7561 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7621 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7681 cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
