LOCUS       sequence-54312-e        6264 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 6264)
  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 6264)
  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-6.20
FEATURES             Location/Qualifiers
     source          1..6264
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        48..244
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      95..230
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      274..283
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             280..1929
                     /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(346..364)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     CDS             1945..2001
                     /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             2002..2937
                     /codon_start=1
                     /product="luciferase from the anthozoan coelenterate 
                     Renilla reniformis (sea pansy)"
                     /label=Rluc
                     /translation="MTSKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    2981..3102
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2992..3011)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3046..3065
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3424..3444
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     primer_bind     complement(3596..3613)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3767..4355)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3847..3866)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4526..5317,5318..5386))
                     /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     5149..5168
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5387..5491)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5518..5973
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5605..5624)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5815..5836
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    6104..6152
                     /note="synthetic polyadenylation signal"
     misc_feature    6166..6257
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     6206..6225
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ggtaccgagc tcttacgcgt gctagcccgg gctcgagatc tgcgatctgc atctcaatta
       61 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
      121 cgcccattct ccgccccatc gctgactaat tttttttatt tatgcagagg ccgaggccgc
      181 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
      241 caaaaagctt ggcattccgg tactgttggt aaagccacca tggaagacgc caaaaacata
      301 aagaaaggcc cggcgccatt ctatccgctg gaagatggaa ccgctggaga gcaactgcat
      361 aaggctatga agagatacgc cctggttcct ggaacaattg cttttacaga tgcacatatc
      421 gaggtggaca tcacttacgc tgagtacttc gaaatgtccg ttcggttggc agaagctatg
      481 aaacgatatg ggctgaatac aaatcacaga atcgtcgtat gcagtgaaaa ctctcttcaa
      541 ttctttatgc cggtgttggg cgcgttattt atcggagttg cagttgcgcc cgcgaacgac
      601 atttataatg aacgtgaatt gctcaacagt atgggcattt cgcagcctac cgtggtgttc
      661 gtttccaaaa aggggttgca aaaaattttg aacgtgcaaa aaaagctccc aatcatccaa
      721 aaaattatta tcatggattc taaaacggat taccagggat ttcagtcgat gtacacgttc
      781 gtcacatctc atctacctcc cggttttaat gaatacgatt ttgtgccaga gtccttcgat
      841 agggacaaga caattgcact gatcatgaac tcctctggat ctactggtct gcctaaaggt
      901 gtcgctctgc ctcatagaac tgcctgcgtg agattctcgc atgccagaga tcctattttt
      961 ggcaatcaaa tcattccgga tactgcgatt ttaagtgttg ttccattcca tcacggtttt
     1021 ggaatgttta ctacactcgg atatttgata tgtggatttc gagtcgtctt aatgtataga
     1081 tttgaagaag agctgtttct gaggagcctt caggattaca agattcaaag tgcgctgctg
     1141 gtgccaaccc tattctcctt cttcgccaaa agcactctga ttgacaaata cgatttatct
     1201 aatttacacg aaattgcttc tggtggcgct cccctctcta aggaagtcgg ggaagcggtt
     1261 gccaagaggt tccatctgcc aggtatcagg caaggatatg ggctcactga gactacatca
     1321 gctattctga ttacacccga gggggatgat aaaccgggcg cggtcggtaa agttgttcca
     1381 ttttttgaag cgaaggttgt ggatctggat accgggaaaa cgctgggcgt taatcaaaga
     1441 ggcgaactgt gtgtgagagg tcctatgatt atgtccggtt atgtaaacaa tccggaagcg
     1501 accaacgcct tgattgacaa ggatggatgg ctacattctg gagacatagc ttactgggac
     1561 gaagacgaac acttcttcat cgttgaccgc ctgaagtctc tgattaagta caaaggctat
     1621 caggtggctc ccgctgaatt ggaatccatc ttgctccaac accccaacat cttcgacgca
     1681 ggtgtcgcag gtcttcccga cgatgacgcc ggtgaacttc ccgccgccgt tgttgttttg
     1741 gagcacggaa agacgatgac ggaaaaagag atcgtggatt acgtcgccag tcaagtaaca
     1801 accgcgaaaa agttgcgcgg aggagttgtg tttgtggacg aagtaccgaa aggtcttacc
     1861 ggaaaactcg acgcaagaaa aatcagagag atcctcataa aggccaagaa gggcggaaag
     1921 atcgccgtgg aattcggaag cggagctact aacttcagcc tgctgaagca ggctggagac
     1981 gtggaggaga accctggacc tatgacttcg aaagtttatg atccagaaca aaggaaacgg
     2041 atgataactg gtccgcagtg gtgggccaga tgtaaacaaa tgaatgttct tgattcattt
     2101 attaattatt atgattcaga aaaacatgca gaaaatgctg ttattttttt acatggtaac
     2161 gcggcctctt cttatttatg gcgacatgtt gtgccacata ttgagccagt agcgcggtgt
     2221 attataccag accttattgg tatgggcaaa tcaggcaaat ctggtaatgg ttcttatagg
     2281 ttacttgatc attacaaata tcttactgca tggtttgaac ttcttaattt accaaagaag
     2341 atcatttttg tcggccatga ttggggtgct tgtttggcat ttcattatag ctatgagcat
     2401 caagataaga tcaaagcaat agttcacgct gaaagtgtag tagatgtgat tgaatcatgg
     2461 gatgaatggc ctgatattga agaagatatt gcgttgatca aatctgaaga aggagaaaaa
     2521 atggttttgg agaataactt cttcgtggaa accatgttgc catcaaaaat catgagaaag
     2581 ttagaaccag aagaatttgc agcatatctt gaaccattca aagagaaagg tgaagttcgt
     2641 cgtccaacat tatcatggcc tcgtgaaatc ccgttagtaa aaggtggtaa acctgacgtt
     2701 gtacaaattg ttaggaatta taatgcttat ctacgtgcaa gtgatgattt accaaaaatg
     2761 tttattgaat cggacccagg attcttttcc aatgctattg ttgaaggtgc caagaagttt
     2821 cctaatactg aatttgtcaa agtaaaaggt cttcattttt cgcaagaaga tgcacctgat
     2881 gaaatgggaa aatatatcaa atcgttcgtt gagcgagttc tcaaaaatga acaataagaa
     2941 ttctagagtc ggggcggccg gccgcttcga gcagacatga taagatacat tgatgagttt
     3001 ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct
     3061 attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt
     3121 cattttatgt ttcaggttca gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc
     3181 tacaaatgtg gtaaaatcga taaggatctg aacgatggag cggagaatgg gcggaactgg
     3241 gcggagttag gggcgggatg ggcggagtta ggggcgggac tatggttgct gactaattga
     3301 gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca cacctggttg
     3361 ctgactaatt gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc
     3421 cacaccctaa ctgacacaca ttccacagcg gatccgtcga ccgatgccct tgagagcctt
     3481 caacccagtc agctccttcc ggtgggcgcg gggcatgact atcgtcgccg cacttatgac
     3541 tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca gcgctcttcc gcttcctcgc
     3601 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     3661 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     3721 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     3781 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     3841 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     3901 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     3961 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     4021 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     4081 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4141 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4201 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4261 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4321 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     4381 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     4441 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     4501 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     4561 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     4621 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     4681 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     4741 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     4801 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     4861 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     4921 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     4981 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     5041 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     5101 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     5161 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     5221 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     5281 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     5341 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     5401 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     5461 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     5521 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     5581 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     5641 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     5701 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     5761 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     5821 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     5881 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     5941 cgaattttaa caaaatatta acgcttacaa tttgccattc gccattcagg ctgcgcaact
     6001 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagcccaag ctaccatgat
     6061 aagtaagtaa tattaaggta cgggaggtac ttggagcggc cgcaataaaa tatctttatt
     6121 ttcattacat ctgtgtgttg gttttttgtg tgaatcgata gtactaacat acgctctcca
     6181 tcaaaacaaa acgaaacaaa acaaactagc aaaataggct gtccccagtg caagtgcagg
     6241 tgccagaaca tttctctatc gata
//
