LOCUS       sequence-339082-        8141 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Accessory Plasmid for PACE of pylBCD variants.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8141)
  AUTHORS   Ho JML, Miller CA, Smith KA, Mattia JR, Bennett MR
  TITLE     Improved pyrrolysine biosynthesis through phage assisted 
            non-continuous directed evolution of the complete pathway.
  JOURNAL   Nat Commun. 2021 Jun 24;12(1):3914. doi: 10.1038/s41467-021-24183-9.
  PUBMED    34168131
REFERENCE   2  (bases 1 to 8141)
  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: "2021-06-24"; volume: "12(1):3914. doi"; pages: " 
            10.1038/s41467-021-24183-9"
COMMENT     Sequence Label: pJH202
FEATURES             Location/Qualifiers
     source          1..8141
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(354..1433)
                     /codon_start=1
                     /gene="luxA"
                     /product="LuxA luciferase subunit"
                     /label=LuxA
                     /note="from the Photorhabdus luminescens bioluminescence 
                     operon"
                     /translation="MKFGNFLLTYQPPQFSQTEVMKRLVKLGRISEECGFDTVWLLEHH
                     FTEFGLLGNPYVAAAYLLGATKKLNVGTAAIVLPTAHPVRQLEDVNLLDQMSKGRFRFG
                     ICRGLYNKDFRVFGTDMNNSRALAECWYGLIKNGMTEGYMEADNEHIKFHKVKVNPAAY
                     SRGGAPVYVVAESASTTEWAAQFGLPMILSWIINTNEKKAQLELYNEVAQEYGHDIHNI
                     DHCLSYITSVDHDSIKAKEICRKFLGHWYDSYVNATTIFDDSDQTRGYDFNKGQWRDFV
                     LKGHKDTNRRIDYSYEINPVGTPQECIDIIQKDIDATGISNICCGFEANGTVDEIIASM
                     KLFQSDVMPFLKEKQRSLLY"
     CDS             3082..3273
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3267..3289)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3375..3515
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3430..3449
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3530..3547)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3701..4289)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3781..3800)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        4350..4421
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4422..5213
                     /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"
     CDS             complement(join(7451..8141,1..290))
                     /codon_start=1
                     /gene="luxB"
                     /product="LuxB luciferase subunit"
                     /label=LuxB
                     /note="from the Photorhabdus luminescens bioluminescence 
                     operon"
                     /translation="KFGLFFLNFINSTTVQEQSIVRMQEITEYVDKLNFEQILVYENHF
                     SDNGVVGAPLTVSGFLLGLTEKIKIGSLNHIITTHHPVRIAEEACLLDQLSEGRFILGF
                     SDCEKKDEMHFFNRPVEYQQQLFEECYEIINDALTTGYCNPDNDFYSFPKISVNPHAYT
                     PGGPRKYVTATSHHIVEWAAKKGIPLIFKWDDSNDVRYEYAERYKAVADKYDVDLSEID
                     HQLMILVNYNEDSNKAKQETRAFISDYVLEMHPNENFENKLEEIIAENAVGNYTECITA
                     AKLAIEKCGAKSVLLSFEPMNDLMSQKNVINIVDDNIKKYHTEYT"
ORIGIN
        1 tcacttaact gatccaataa gcaagcttcc tccgctatgc ggacaggatg atgagttgta
       61 atgatgtgat ttaatgaacc aattttaatt ttctctgtta aaccgagcag aaaaccagaa
      121 acagtcagag gagcgccgac aacaccatta tctgaaaaat gattttcata cactaaaatc
      181 tgttcaaaat tcaacttatc aacatactcc gttatttcct gcatgcgaac tatactctgt
      241 tcttgaacag ttgttgaatt gatgaagtta aggaagaaca atccaaattt gctaccgcca
      301 ccgccgctac cgccaccgcc gctaccgcca ccgccgctac cgccaccgcc ataatataat
      361 agcgaacgtt gtttttcttt aagaaatggc atgacatcag actggaagag cttcatggaa
      421 gcaataattt cgtctactgt tccattagct tcaaatccac aacaaatatt tgatattcct
      481 gtagcatcaa tgtctttttg aattatgtca atacattcct gcggcgttcc cacgggattg
      541 atttcgtaac tgtaatcaat acggcgatta gtatctttat gtccttttaa tacaaagtca
      601 cgccactgcc ctttattgaa atcataacct cttgtttggt ctgaatcatc aaaaatagtc
      661 gtagcattca cataagaatc ataccaatgc cccagaaatt tccggcaaat ctctttcgct
      721 ttaattgagt catgatctac agatgttata tatgataagc aatggtcgat attatgaata
      781 tcgtgcccat attcttgagc cacttcatta taaagctcaa gttgtgcttt cttttcgtta
      841 gtatttataa tccaacttaa tatcatcggt aggccaaatt gagcagccca ctcagtcgtc
      901 gaagctgatt cagccaccac ataaaccggt gcgccacctc tgctatacgc cgcggggttt
      961 acttttacct tatggaactt gatatgttca ttatcagctt ccatatatcc ctctgtcatg
     1021 ccattcttta tcagcccgta ccagcattcc gctaaggcgc gactgttatt catatctgtg
     1081 ccgaatacgc gaaagtcctt gttgtaaagc cctcggcaaa taccaaaccg aaatcgtcct
     1141 tttgacattt gatccaataa attcacatct tcaagttggc gtactggatg ggctgtggga
     1201 agaacaatag cggcagttcc tacattcaat tttttagtcg cgccaagtaa atatgcagca
     1261 gcgacataag ggttaccaag caaaccaaac tccgtgaaat gatgctccag taaccatacg
     1321 gtatcaaaac cacactcctc agagatgcga cctaatttaa ccaaacgttt cattacctct
     1381 gtttgggaaa attggggagg ttggtatgta agcaaaaagt ttccaaattt cattaagact
     1441 ccttattacg cagtatgtta gcaaacgtag aaaatacata cataaaggtg gcaacatata
     1501 aaagaaacgc aaagacaccg cggaacaggt tgatcttatc gcagtcgata ctgaactcgt
     1561 aaggtttacc agcgccaaag acaaaagggc gacattcaac cgattgaggg agggaaggta
     1621 aatattgacg gaaattattc attaaaggtg aattatcacc gtcaccgact tgagccattt
     1681 gggaattaga gccagcaaaa tcaccagtag caccattacc attagcaagg ccggaaacgt
     1741 caccaatgaa accatcgata gcagcaccgt aatcagtagc gacagaatca agtttgcctt
     1801 tagcgtcaga ctgtagcgcg ttttcatcgg cattttcggt catagccccc ttattagcgt
     1861 ttgccatctt ttcataatca aaatcaccgg aaccagagcc accaccggaa ccgcctccct
     1921 cagagccgcc accctcagaa ccgccaccct cagagccacc accctcagag ccgccaccag
     1981 aaccaccacc agagccgccg ccagcattga caggaggttg aggcaggtca gacgattggc
     2041 cttgatattc acaaacgaat ggatcctcat taaagccaga atggaaagcg cagtctctga
     2101 atttaccgtt ccagtaagcg tcatacatgg cttttgatga tacaggagtg tactggtaat
     2161 aagttttaac ggggtcagtg ccttgagtaa cagtgcccgt ataaacagtt aatgccccct
     2221 gcctatttcg gaacctatta ttctgaaaca tgaaagtatt aagaggctga gactcctcaa
     2281 gagaaggatt aggattagcg gggttttgct cagtaccagg cggataagtg ccgtcgagag
     2341 ggttgatata agtatagccc ggaataggtg tatcaccgta ctcaggaggt ttagtaccgc
     2401 caccctcaga accgccaccc tcagaaccgc caccctcaga gccaccaccc tcattttcag
     2461 ggatagcaag cccaatagga acccatgtac cgtaacactg agtttcgtca ccagtacaaa
     2521 ctacaacgcc tgtagcattc cacagacagc cctcatagtt agcgtaacga tctaaagttt
     2581 tgtcgtcttt ccagacgtta gtaaatgaat tttctgtatg ctattttgct aaacaacttt
     2641 caacagtttc agcggagtga gaatagaaag gaacaactaa aggaattgcg aataataatt
     2701 ttttcatttt tttttcctcc ttactgcacc tgcaggtaat gttgtcctct tgatttctgc
     2761 gttcaggatt gtcctgctga actgttaata caatttgcgt gccaattttt tatctttttg
     2821 atttataaag atctgattga agaatcaaca gcaacatgcc aggatgagtt agcgaattac
     2881 actaacaagt ggcgaatttc atcagggtcc ctaagtctcc tcagcaaaac gaaaggccca
     2941 gtctttcgac tgagcctttc gttttatttg accggatgtc ctctttgttc atcatcagta
     3001 acccgtatcg tgagcatcct ctctcgtttc atcggtatca ttacccccat gaacagaaat
     3061 cccccttaca cggaggcatc agtgaccaaa caggaaaaaa ccgcccttaa catggcccgc
     3121 tttatcagaa gccagacatt aacgcttctg gagaaactca acgagctgga cgcggatgaa
     3181 caggcagaca tctgtgaatc gcttcacgac cacgctgatg agctttaccg cagctgcctc
     3241 gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca
     3301 gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt
     3361 ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc
     3421 ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac
     3481 cgcacagatg cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg
     3541 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     3601 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     3661 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     3721 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     3781 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     3841 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     3901 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     3961 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     4021 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     4081 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     4141 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agaggtggta
     4201 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     4261 agattacgcg cagaaaaaaa ggatctcaac ggcctatttg gcctggaaag cgggcagtga
     4321 gcgcaacgca attaatattt ttctaaatac attcaaatat gtatacgctc atgaaacaat
     4381 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagggaa gcggtgatcg
     4441 ccgaagtatc gactcaacta tcagaggtag ttggcgtcat cgagcgccat ctcgaaccga
     4501 cgttgctggc cgtacatttg tacggctccg cagtggatgg cggcctgaag ccacacagtg
     4561 atattgattt gctggttacg gtgaccgtaa ggcttgatga aacaacgcgg cgagctttga
     4621 tcaacgacct tttggaaact tcggcttccc ctggagagag cgagattctc cgcgctgtag
     4681 aagtcaccat tgttgtgcac gacgacatca ttccgtggcg ttatccagct aagcgcgaac
     4741 tgcaatttgg agaatggcag cgcaatgaca ttcttgcagg tatcttcgag ccagccacga
     4801 tcgacattga tctggctatc ttgctgacaa aagcaagaga acatagcgtt gccttggtag
     4861 gtccagcggc ggaggaactc tttgatccgg ttcctgaaca ggatctattt gaggcgctaa
     4921 atgaaacctt aacgctatgg aactcgccgc ccgactgggc tggcgatgag cgaaatgtag
     4981 tgcttacgtt gtcccgcatt tggtacagcg cagtaaccgg caaaatcgcg ccgaaggatg
     5041 tcgctgccga ctgggcaatg gagcgcctgc cggcccagta tcagcccgtc atacttgaag
     5101 ctagacaggc ttatcttgga caagaagaag atcgcttggc ctcgcgcgca gatcagttgg
     5161 aagaatttgt ccactacgtg aaaggcgaga tcaccaaggt agtcggcaaa taaacgccat
     5221 gggcattaaa aaatgccctc ttgggttatc aagagggtca ttatatttgc tgaggagtgc
     5281 cgttaattaa gtccatcatg attacgccaa gcttgcatgc aggcctctgc agtcgacggg
     5341 cccgggatcc gatgaacgtc ggcagaatgc ttaatgaatt acaacagtac tgcgatgagt
     5401 ggcagggcgg ggcgtaattt ttttaaggca gttattggtg cccttaaacg cctggttgct
     5461 acgcctgaat aagtgataat aagcggatga atggcagaag cttaaaaaaa atccttagct
     5521 ttcgctaagg atctgcaggt cgacggatcc tggcggaaac cccgggaatc taacccggct
     5581 gaacggattt agagtccatt cgatctacat gatcaggttt ccgaatgcgg ggcgcatctt
     5641 actgcgcaga tacgccctcg tcaatccctt aatagcaaaa tgccttttga tcggcgagaa
     5701 agtcagcagg ccgcttagtt agccgtgcac tgatcatatc gtcaattatt acctccacgg
     5761 ggagagcctg agcaaactgg cctcaggcat ttgagaagca cacggacagt catttgatcg
     5821 gcgagaaagt cagcaggccg cttagttagc cagggtggtt tttcttttca ccagcgagac
     5881 gggcaacaga acgccatgag cggcctcatt tcttattatg agttacaaca gtccgcaccg
     5941 ctgccggtag ctccttcctg tgggcgcggg ccatagatct cagcggtggc agcagcctag
     6001 gttacaggtt ggtagaaatc ccgttatagt aagactcgga ccttgcagct ctcttgatat
     6061 ttttaaagtc gtgtttaacc tttagaaggc gttcgagtcc gaaacctgcc cctatccagg
     6121 gtttatcaat accccattcc cggtcaagcg gtatgggtcc gactactgca gaggaaagtt
     6181 ccaggtctcc gtgcattaca tcaagggtat ccccaaagac catgcaggaa tcgcctacga
     6241 tcttgaaatc aattcccagg tggttcagga agtccgtaat tatgctttca agattttccc
     6301 gtgtgcatcc cgatcccatc tggcagaagt tcagcatggt aaactcttcg aggtgttctt
     6361 tgccgtcgga ctcttttctg tagcatgggc ctatttcaaa aatttttatt ggatcaggca
     6421 gggccctgtc aagcttgcgc aggtagttgt aaaggtttgg agcaagcatg ggtctcaggc
     6481 agaagttctg gtcaaccctg aagatctgtt ttgaaagttc ggtatcatta tcaatgccca
     6541 tcctttcgat atactcaaga gggatcagga tcggggattt tatttccaga aaacccctgt
     6601 ccacaaagaa cctggtaatt tcacgctcga gtttccccag ataattctcc ctttcttccg
     6661 cgtagatctg ctgcaggtct ttttttctgc gagagagcaa ttcggactca agctccctga
     6721 aaggcttgcc ggaattcagg gaaatctcat cttttgggtt taacaggact tcaagcctgt
     6781 cagtctggct cttcgtaagt gcgggggcac ttgcagaggt cggaaccgga gagttcgggg
     6841 tagatttcgc cggagacgga acagaacgag aggtgtcggt agacgctttc gcagaaaccg
     6901 gatttttcag cggtttcggc gcacgagaaa cagatttcgg catcgctttt ttcactttcg
     6961 gcgcagaaac aactttaact ttaacagagg ttttgccttc ggtagaacgg gtcaggaagt
     7021 tgttgatatc ttcgtcagaa acacgacaac gtttgcaggt tttacggtat ttatgataac
     7081 ggaatgcacg tgcgggacgg caagagcgag agttgttcac aaccagatgg tcaccacacg
     7141 ccatttcgat gtagatttta gaacgagaaa cctcatagtg cttgatcttg tgcagcgtgc
     7201 cggtacggga catccacaga ccggtcgcag agatcagaac atccagcggc ttattattca
     7261 tactagttac cctctagatt gagttaatct ccatgtagcg ttacaagtat tacacaaagt
     7321 tttttatgtt gagaatattt ttttgatggg aaggcactta tttttgatcg ttcgctcaaa
     7381 gaagcatcga acgtagggcc ctaaaaaaaa accccgctga agcggggttt tttgcgtaca
     7441 ttggctccgt ttaggtatat tccgtgtggt acttcttaat attatcatca acaatattga
     7501 ttacattttt ttggctcatc aaatcattca ttggttcaaa ggacagcaat acacttttcg
     7561 caccacactt ttcaattgcc aacttagccg cagttataca ctccgtataa tttccgacag
     7621 cgttttctgc aattatttct tcaagtttat tttcgaaatt ttcattaggg tgcatttcaa
     7681 gaacataatc actaataaat gcacgcgtct cttgtttagc tttattacta tcttcgttat
     7741 agttaactaa tatcattaac tgatggtcta tctctgatag gtcaacgtca tatttatccg
     7801 caacggcttt atatctttca gcatattcat atctaacatc attagaatca tcccacttaa
     7861 agatgagagg aatacctttt ttggccgccc actcaacaat atgatgactg gttgctgtta
     7921 catatttccg aggtccgcct ggcgtataag catggggatt tacagatatt ttagggaagc
     7981 tataaaaatc gttatctgga ttacaatagc ctgttgttaa agcatcgtta atgatttcat
     8041 aacactcttc aaatagttgc tgttgatatt caaccgggcg attaaaaaaa tgcatttcat
     8101 cttttttttc gcaatcacta aaccctaaaa taaatctccc t
//
