LOCUS       sequence-300360-        6539 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses orthogonal firefly luciferase gene in bacterial cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6539)
  AUTHORS   Aleksashin NA, Szal T, d'Aquino AE, Jewett MC, Vazquez-Laslop N, 
            Mankin AS
  TITLE     A fully orthogonal system for protein synthesis in bacterial cells.
  JOURNAL   Nat Commun. 2020 Apr 20;11(1):1858. doi: 10.1038/s41467-020-15756-1.
  PUBMED    32313034
REFERENCE   2  (bases 1 to 6539)
  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 
            Commun."; date: "2020-04-20"; volume: "11(1):1858. doi"; pages: " 
            10.1038/s41467-020-15756-1"
COMMENT     Sequence Label: poLuc1
FEATURES             Location/Qualifiers
     source          1..6539
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    21..39
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    46..64
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             83..94
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      898..969
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      985..1012
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             1114..2766
                     /codon_start=1
                     /gene="luc"
                     /product="firefly luciferase"
                     /label=luciferase
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKSKL"
     primer_bind     complement(1180..1198)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     primer_bind     complement(2836..2853)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      3006..3092
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3184..3211
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        complement(join(3657..3662,3663..3679,3680..3685))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     rep_origin      3797..4342
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     4243..4262
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4459..4476
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             5530..6321
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
ORIGIN
        1 attcgcggcc gcttctagag tccctatcag tgatagagat tgacatccct atcagtgata
       61 gagatactga gcactactag agaaagagga gaaatactag atgaaaaaca taaatgccga
      121 cgacacatac agaataatta ataaaattaa agcttgtaga agcaataatg atattaatca
      181 atgcttatct gatatgacta aaatggtaca ttgtgaatat tatttactcg cgatcattta
      241 tcctcattct atggttaaat ctgatatttc aatcctagat aattacccta aaaaatggag
      301 gcaatattat gatgacgcta atttaataaa atatgatcct atagtagatt attctaactc
      361 caatcattca ccaattaatt ggaatatatt tgaaaacaat gctgtaaata aaaaatctcc
      421 aaatgtaatt aaagaagcga aaacatcagg tcttatcact gggtttagtt tccctattca
      481 tacggctaac aatggcttcg gaatgcttag ttttgcacat tcagaaaaag acaactatat
      541 agatagttta tttttacatg cgtgtatgaa cataccatta attgttcctt ctctagttga
      601 taattatcga aaaataaata tagcaaataa taaatcaaac aacgatttaa ccaaaagaga
      661 aaaagaatgt ttagcgtggg catgcgaagg aaaaagctct tgggatattt caaaaatatt
      721 aggttgcagt gagcgtactg tcactttcca tttaaccaat gcgcaaatga aactcaatac
      781 aacaaaccgc tgccaaagta tttctaaagc aattttaaca ggagcaattg attgcccata
      841 ctttaaaaat taataacact gatagtgcta gtgtagatca ctactagagc caggcatcaa
      901 ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg
      961 aacgctctct actagagtca cactggctca ccttcgggtg ggcctttctg cgtttatata
     1021 ctagagacct gtaggatcgt acaggtttac gcaagaaaat ggtttgttat agtcgaataa
     1081 actatatctg ttattttttc caaccacaga tctatggaag acgccaaaaa cataaagaaa
     1141 ggcccggcgc cattctatcc gctagaggat ggaaccgctg gagagcaact gcataaggct
     1201 atgaagagat acgccctggt tcctggaaca attgctttta cagatgcaca tatcgaggtg
     1261 aacatcacgt acgcggaata cttcgaaatg tccgttcggt tggcagaagc tatgaaacga
     1321 tatgggctga atacaaatca cagaatcgtc gtatgcagtg aaaactctct tcaattcttt
     1381 atgccggtgt tgggcgcgtt atttatcgga gttgcagttg cgcccgcgaa cgacatttat
     1441 aatgaacgtg aattgctcaa cagtatgaac atttcgcagc ctaccgtagt gtttgtttcc
     1501 aaaaaggggt tgcaaaaaat tttgaacgtg caaaaaaaat taccaataat ccagaaaatt
     1561 attatcatgg attctaaaac ggattaccag ggatttcagt cgatgtacac gttcgtcaca
     1621 tctcatctac ctcccggttt taatgaatac gattttgtac cagagtcctt tgatcgtgac
     1681 aaaacaattg cactgataat gaactcctct ggatctactg ggttacctaa gggtgtggcc
     1741 cttccgcata gaactgcctg cgtcagattc tcgcatgcca gagatcctat ttttggcaat
     1801 caaatcattc cggatactgc gattttaagt gttgttccat tccatcacgg ttttggaatg
     1861 tttactacac tcggatattt gatatgtgga tttcgagtcg tcttaatgta tagatttgaa
     1921 gaagagctgt ttttacgatc ccttcaggat tacaaaattc aaagtgcgtt gctagtacca
     1981 accctatttt cattcttcgc caaaagcact ctgattgaca aatacgattt atctaattta
     2041 cacgaaattg cttctggggg cgcacctctt tcgaaagaag tcggggaagc ggttgcaaaa
     2101 cgcttccatc ttccagggat acgacaagga tatgggctca ctgagactac atcagctatt
     2161 ctgattacac ccgaggggga tgataaaccg ggcgcggtcg gtaaagttgt tccatttttt
     2221 gaagcgaagg ttgtggatct ggataccggg aaaacgctgg gcgttaatca gagaggcgaa
     2281 ttatgtgtca gaggacctat gattatgtcc ggttatgtaa acaatccgga agcgaccaac
     2341 gccttgattg acaaggatgg atggctacat tctggagaca tagcttactg ggacgaagac
     2401 gaacacttct tcatagttga ccgcttgaag tctttaatta aatacaaagg ataccaggtg
     2461 gcccccgctg aattggagtc gatattgtta caacacccca acatcttcga cgcgggcgtg
     2521 gcaggtcttc ccgacgatga cgccggtgaa cttcccgccg ccgttgttgt tttggagcac
     2581 ggaaagacga tgacggaaaa agagatcgtg gattacgtcg ccagtcaagt aacaaccgcc
     2641 aaaaagttgc gcggaggagt tgtgtttgtg gacgaagtac cgaaaggtct taccggaaaa
     2701 ctcgacgcaa gaaaaatcag agagatcctc ataaaggcca agaagggcgg aaagtccaaa
     2761 ttgtaagtcg accggctgct aacaaagccc gcggccgctg aagatcgatc tcgacgagtg
     2821 agagaagatt ttcagcctga tacagattaa atcagaacgc agaagcggtc tgataaaaca
     2881 gaatttgcct ggcggcagta gcgcggtggt cccacctgac cccatgccga actcagaagt
     2941 gaaacgccgt agcgccgatg gtagtgtggg gtcaccccat gcgagagtag ggaactgcca
     3001 ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt
     3061 tgtcggtgaa cgctctcctg agtaggacaa atccgccggg agcggatttg aacgttgcga
     3121 agcaacggcc cggagggtgg cgggcaggac gcccgccata aactgccagg catcaaatta
     3181 agcagaaggc catcctgacg gatggccttt ttgcgtttct acactcgagg atctctttcc
     3241 tttccatcaa aaaaatattg atgaaatgag ctgtggggcg ttatttaggt tttttcttct
     3301 ttcgaaaaaa tctttcttta tgaagttaaa agctatgtat tcaatagcat attttgaata
     3361 tggacataga atagtgctta tcactattgc atatagcatc ttatctgaca caaggaaata
     3421 atacccttcg ctgttttttg ttataaggta tatatatata agtgtgcagt acaggccaaa
     3481 taaaatattt tttatgtagt atcttaaatc ccgcaagagg cccggcagta ccggcataac
     3541 caagcctatg cctacagcat ccagggtgac ggtgccgagg atgacgatga gcgcattgtt
     3601 agatttcata cacggtgcct gactgcgtta gcaatttaac tgtgataaac taccgcatta
     3661 aagcttatcg atgataagct gtcaaacatg agaattacaa cttatatcgt atggggctga
     3721 cttcaggtgc tacatttgaa gagataaatt gcactgaaat ctagaaatat tttatctgat
     3781 taataagatg atcttcttga gatcgttttg gtctgcgcgt aatctcttgc tctgaaaacg
     3841 aaaaaaccgc cttgcagggc ggtttttcga aggttctctg agctaccaac tctttgaacc
     3901 gaggtaactg gcttggagga gcgcagtcac caaaacttgt cctttcagtt tagccttaac
     3961 cggcgcatga cttcaagact aactcctcta aatcaattac cagtggctgc tgccagtggt
     4021 gcttttgcat gtctttccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     4081 tcggactgaa cggggggttc gtgcatacag tccagcttgg agcgaactgc ctacccggaa
     4141 ctgagtgtca ggcgtggaat gagacaaacg cggccataac agcggaatga caccggtaaa
     4201 ccgaaaggca ggaacaggag agcgcacgag ggagccgcca gggggaaacg cctggtatct
     4261 ttatagtcct gtcgggtttc gccaccactg atttgagcgt cagatttcgt gatgcttgtc
     4321 aggggggcgg agcctatgga aaaacggctt tgccgcggcc ctctcacttc cctgttaagt
     4381 atcttcctgg catcttccag gaaatctccg ccccgttcgt aagccatttc cgctcgccgc
     4441 agtcgaacga ccgagcgtag cgagtcagtg agcgaggaag cggaatatat cctgtatcac
     4501 atattctgct gacgcaccgg tgcagccttt tttctcctgc cacatgaagc acttcactga
     4561 caccctcatc agtgccaaca tagtaagcca gtatacactc cgctagcgct gatgtccggc
     4621 ggtgcttttg ccgttacgca ccaccccgtc agtagctgaa caggagggac agagctttat
     4681 gcttgtaaac cgttttgtga aaaaattttt aaaataaaaa aggggacctc tagggtcccc
     4741 aattaattag taatataatc tattaaaggt cattcaaaag gtcatccacc ggatcaattc
     4801 ccctgctcgc gcaggctggg tgccaagctc tcgggtaaca tcaaggcccg atccttggag
     4861 cccttgccct cccgcacgat gatcgtgccg tgatcgaaaa tccagatcct tgacccgcat
     4921 ttgcaaaccc tcactgatcc gcatggggcc cccgttccat acagaagctg ggcgaacaaa
     4981 cgatgctcgc cttccagaaa accgaggatg cgaaccactt catccggggt cagcaccacc
     5041 ggcaagcgcc gcgacggccg aggtcttccg atctcctgaa gccagggcag atccgtgcac
     5101 agcaccttgc cgtagaagaa cagcaaggcc gccaatgcct gacgatgcgt ggagaccgaa
     5161 accttgcgct cgttcgccag ccaggacaga aatgcctcga cttcgctgct gcccaaggtt
     5221 gccgggtgac gcacaccgtg gaaacggatg aaggcacgaa cccagtggac ataagcctgt
     5281 tcggttcgta agctgtaatg caagtagcgt atgcgctcac gcaactggtc cagaaccttg
     5341 accgaacgca gcggtggtaa cggcgcagtg gcggttttca tggcttgtta tgactgtttt
     5401 tttggggtac agtctatgcc tcgggcatcc aagcagcaag cgcgttacgc cgtgggtcga
     5461 tgtttgatgt tatggagcag caacgatgtt acgcagcagg gcagtcgccc taaaacaaag
     5521 ttaaacatca tgagggaagc ggtgatcgcc gaagtatcga ctcaactatc agaggtagtt
     5581 ggcgtcatcg agcgccatct cgaaccgacg ttgctggccg tacatttgta cggctccgca
     5641 gtggatggcg gcctgaagcc acacagtgat attgatttgc tggttacggt gaccgtaagg
     5701 cttgatgaaa caacgcggcg agctttgatc aacgaccttt tggaaacttc ggcttcccct
     5761 ggagagagcg agattctccg cgctgtagaa gtcaccattg ttgtgcacga cgacatcatt
     5821 ccgtggcgtt atccagctaa gcgcgaactg caatttggag aatggcagcg caatgacatt
     5881 cttgcaggta tcttcgagcc agccacgatc gacattgatc tggctatctt gctgacaaaa
     5941 gcaagagaac atagcgttgc cttggtaggt ccagcggcgg aggaactctt tgatccggtt
     6001 cctgaacagg atctatttga ggcgctaaat gaaaccttaa cgctatggaa ctcgccgccc
     6061 gactgggctg gcgatgagcg aaatgtagtg cttacgttgt cccgcatttg gtacagcgca
     6121 gtaaccggca aaatcgcgcc gaaggatgtc gctgccgact gggcaatgga gcgcctgccg
     6181 gcccagtatc agcccgtcat acttgaagct agacaggctt atcttggaca agaagaagat
     6241 cgcttggcct cgcgcgcaga tcagttggaa gaatttgtcc actacgtgaa aggcgagatc
     6301 accaaggtag tcggcaaata atgtctaaca attcgttcaa gccgacgccg cttcgcggcg
     6361 cggcttaact caagcgttag atgcactaag cacataattg ctcacagcca aactatcagg
     6421 tcaagtctgc ttttattatt tttaagcgtg cataataagc cctacacaaa ttgggagata
     6481 tatcatgaaa ggctggcttt ttcttgttat cgcaatagtt ggcgaagtaa tcgcaacat
//
