LOCUS       sequence-54359-e        6206 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6206)
  AUTHORS   Cheng KC, Inglese J
  TITLE     A coincidence reporter-gene system for high-throughput screening.
  JOURNAL   Nat Methods. 2012 Oct;9(10):937. doi: 10.1038/nmeth.2170.
  PUBMED    23018994
REFERENCE   2  (bases 1 to 6206)
  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"; journalName: "Nat 
            Methods."; date: "2012-10"; volume: "9(10):937. doi"; pages: " 
            10.1038/nmeth.2170"
COMMENT     Sequence Label: pCI-6.24
FEATURES             Location/Qualifiers
     source          1..6206
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        152..183
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     regulatory      216..225
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             222..1871
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(288..306)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     CDS             1887..1943
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1944..2879
                     /codon_start=1
                     /product="luciferase from the anthozoan coelenterate 
                     Renilla reniformis (sea pansy)"
                     /label=Rluc
                     /translation="MTSKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVVRVLKNEQ"
     polyA_signal    2923..3044
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2934..2953)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2988..3007
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3366..3386
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     primer_bind     complement(3538..3555)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3709..4297)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3789..3808)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4468..5259,5260..5328))
                     /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     5091..5110
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5329..5433)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5460..5915
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5547..5566)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5757..5778
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    6046..6094
                     /note="synthetic polyadenylation signal"
     misc_feature    6108..6199
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     6148..6167
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ggtaccgagc tcttacgcgt gctagcccgg gctcgagatc tgcgatctaa gtaagcttgc
       61 tcgagatctg cgatctaaga gcctgacgtc agagagcctg acgtcagaga gcctgacgtc
      121 agagagcctg acgtcagagg aattcagaca ctagagggta tataatggaa gctcgacttc
      181 cagcttaagc ttggcattcc ggtactgttg gtaaagccac catggaagac gccaaaaaca
      241 taaagaaagg cccggcgcca ttctatccgc tggaagatgg aaccgctgga gagcaactgc
      301 ataaggctat gaagagatac gccctggttc ctggaacaat tgcttttaca gatgcacata
      361 tcgaggtgga catcacttac gctgagtact tcgaaatgtc cgttcggttg gcagaagcta
      421 tgaaacgata tgggctgaat acaaatcaca gaatcgtcgt atgcagtgaa aactctcttc
      481 aattctttat gccggtgttg ggcgcgttat ttatcggagt tgcagttgcg cccgcgaacg
      541 acatttataa tgaacgtgaa ttgctcaaca gtatgggcat ttcgcagcct accgtggtgt
      601 tcgtttccaa aaaggggttg caaaaaattt tgaacgtgca aaaaaagctc ccaatcatcc
      661 aaaaaattat tatcatggat tctaaaacgg attaccaggg atttcagtcg atgtacacgt
      721 tcgtcacatc tcatctacct cccggtttta atgaatacga ttttgtgcca gagtccttcg
      781 atagggacaa gacaattgca ctgatcatga actcctctgg atctactggt ctgcctaaag
      841 gtgtcgctct gcctcataga actgcctgcg tgagattctc gcatgccaga gatcctattt
      901 ttggcaatca aatcattccg gatactgcga ttttaagtgt tgttccattc catcacggtt
      961 ttggaatgtt tactacactc ggatatttga tatgtggatt tcgagtcgtc ttaatgtata
     1021 gatttgaaga agagctgttt ctgaggagcc ttcaggatta caagattcaa agtgcgctgc
     1081 tggtgccaac cctattctcc ttcttcgcca aaagcactct gattgacaaa tacgatttat
     1141 ctaatttaca cgaaattgct tctggtggcg ctcccctctc taaggaagtc ggggaagcgg
     1201 ttgccaagag gttccatctg ccaggtatca ggcaaggata tgggctcact gagactacat
     1261 cagctattct gattacaccc gagggggatg ataaaccggg cgcggtcggt aaagttgttc
     1321 cattttttga agcgaaggtt gtggatctgg ataccgggaa aacgctgggc gttaatcaaa
     1381 gaggcgaact gtgtgtgaga ggtcctatga ttatgtccgg ttatgtaaac aatccggaag
     1441 cgaccaacgc cttgattgac aaggatggat ggctacattc tggagacata gcttactggg
     1501 acgaagacga acacttcttc atcgttgacc gcctgaagtc tctgattaag tacaaaggct
     1561 atcaggtggc tcccgctgaa ttggaatcca tcttgctcca acaccccaac atcttcgacg
     1621 caggtgtcgc aggtcttccc gacgatgacg ccggtgaact tcccgccgcc gttgttgttt
     1681 tggagcacgg aaagacgatg acggaaaaag agatcgtgga ttacgtcgcc agtcaagtaa
     1741 caaccgcgaa aaagttgcgc ggaggagttg tgtttgtgga cgaagtaccg aaaggtctta
     1801 ccggaaaact cgacgcaaga aaaatcagag agatcctcat aaaggccaag aagggcggaa
     1861 agatcgccgt ggaattcgga agcggagcta ctaacttcag cctgctgaag caggctggag
     1921 acgtggagga gaaccctgga cctatgactt cgaaagttta tgatccagaa caaaggaaac
     1981 ggatgataac tggtccgcag tggtgggcca gatgtaaaca aatgaatgtt cttgattcat
     2041 ttattaatta ttatgattca gaaaaacatg cagaaaatgc tgttattttt ttacatggta
     2101 acgcggcctc ttcttattta tggcgacatg ttgtgccaca tattgagcca gtagcgcggt
     2161 gtattatacc agaccttatt ggtatgggca aatcaggcaa atctggtaat ggttcttata
     2221 ggttacttga tcattacaaa tatcttactg catggtttga acttcttaat ttaccaaaga
     2281 agatcatttt tgtcggccat gattggggtg cttgtttggc atttcattat agctatgagc
     2341 atcaagataa gatcaaagca atagttcacg ctgaaagtgt agtagatgtg attgaatcat
     2401 gggatgaatg gcctgatatt gaagaagata ttgcgttgat caaatctgaa gaaggagaaa
     2461 aaatggtttt ggagaataac ttcttcgtgg aaaccatgtt gccatcaaaa atcatgagaa
     2521 agttagaacc agaagaattt gcagcatatc ttgaaccatt caaagagaaa ggtgaagttc
     2581 gtcgtccaac attatcatgg cctcgtgaaa tcccgttagt aaaaggtggt aaacctgacg
     2641 ttgtacaaat tgttaggaat tataatgctt atctacgtgc aagtgatgat ttaccaaaaa
     2701 tgtttattga atcggaccca ggattctttt ccaatgctat tgttgaaggt gccaagaagt
     2761 ttcctaatac tgaatttgtc aaagtaaaag gtcttcattt ttcgcaagaa gatgcacctg
     2821 atgaaatggg aaaatatatc aaatcgttcg ttgtgcgagt tctcaaaaat gaacaataag
     2881 aattctagag tcggggcggc cggccgcttc gagcagacat gataagatac attgatgagt
     2941 ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg
     3001 ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca
     3061 ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc
     3121 tctacaaatg tggtaaaatc gataaggatc tgaacgatgg agcggagaat gggcggaact
     3181 gggcggagtt aggggcggga tgggcggagt taggggcggg actatggttg ctgactaatt
     3241 gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc cacacctggt
     3301 tgctgactaa ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt
     3361 tccacaccct aactgacaca cattccacag cggatccgtc gaccgatgcc cttgagagcc
     3421 ttcaacccag tcagctcctt ccggtgggcg cggggcatga ctatcgtcgc cgcacttatg
     3481 actgtcttct ttatcatgca actcgtagga caggtgccgg cagcgctctt ccgcttcctc
     3541 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     3601 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3661 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     3721 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     3781 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     3841 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3901 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3961 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     4021 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     4081 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     4141 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     4201 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     4261 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     4321 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     4381 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     4441 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     4501 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     4561 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     4621 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     4681 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     4741 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     4801 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     4861 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     4921 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     4981 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     5041 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     5101 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     5161 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     5221 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     5281 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     5341 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     5401 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     5461 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     5521 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     5581 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     5641 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     5701 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     5761 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     5821 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     5881 cgcgaatttt aacaaaatat taacgcttac aatttgccat tcgccattca ggctgcgcaa
     5941 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagccca agctaccatg
     6001 ataagtaagt aatattaagg tacgggaggt acttggagcg gccgcaataa aatatcttta
     6061 ttttcattac atctgtgtgt tggttttttg tgtgaatcga tagtactaac atacgctctc
     6121 catcaaaaca aaacgaaaca aaacaaacta gcaaaatagg ctgtccccag tgcaagtgca
     6181 ggtgccagaa catttctcta tcgata
//
