LOCUS       sequence-192366-        6601 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Accessory plasmid for PACE of PylRS variants.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6601)
  AUTHORS   Bryson DI, Fan C, Guo LT, Miller C, Soll D, Liu DR
  TITLE     Continuous directed evolution of aminoacyl-tRNA synthetases.
  JOURNAL   Nat Chem Biol. 2017 Oct 16. doi: 10.1038/nchembio.2474.
  PUBMED    29035361
REFERENCE   2  (bases 1 to 6601)
  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 Chem 
            Biol. 2017 Oct 16. doi: 10.1038/nchembio.2474."
COMMENT     Sequence Label: pDB038b
FEATURES             Location/Qualifiers
     source          1..6601
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      25..613
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     514..533
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     767..784
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    799..939
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(865..884)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1025..1047
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1041..1232)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             2881..3960
                     /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             4024..5004
                     /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"
     CDS             complement(5702..6493)
                     /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"
     promoter        complement(6494..6565)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccgctttcca ggccaaatag gccgttgaga tccttttttt ctgcgcgtaa tctgctgctt
       61 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccacc
      121 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt
      181 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
      241 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
      301 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
      361 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
      421 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
      481 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
      541 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
      601 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
      661 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
      721 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
      781 cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
      841 acaccgcata tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta
      901 tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc
      961 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     1021 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag
     1081 ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc ctgttcatcc
     1141 gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc
     1201 atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg
     1261 ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat
     1321 gatgaacaaa gaggacatcc ggtcaaataa aacgaaaggc tcagtcgaaa gactgggcct
     1381 ttcgttttgc tgaggagact tagggaccct gatgaaattc gccacttgtt agtgtaattc
     1441 gctaactcat cctggcatgt tgctgttgat tcttcaatca gatctttata aatcaaaaag
     1501 ataaaaaatt ggcacgcaaa ttgtattaac agttcagcag gacaatcctg aacgcagaaa
     1561 tcaagaggac aacattacct gcaggtgcag taaggaggaa aaaaaaatga aaaaattatt
     1621 attcgcaatt cctttagttg ttcctttcta ttctcactcc gctgaaactg ttgaaagttg
     1681 tttagcaaaa tagcatacag aaaattcatt tactaacgtc tggaaagacg acaaaacttt
     1741 agatcgttac gctaactatg agggctgtct gtggaatgct acaggcgttg tagtttgtac
     1801 tggtgacgaa actcagtgtt acggtacatg ggttcctatt gggcttgcta tctaggaaaa
     1861 tgagggtggt ggctctgagg gtggcggttc tgagggtggc ggttctgagg gtggcggtac
     1921 taaacctcct gagtacggtg atacacctat tccgggctat acttatatca accctctcga
     1981 cggcacttat ccgcctggta ctgagcaaaa ccccgctaat cctaatcctt ctcttgagga
     2041 gtctcagcct cttaatactt tcatgtttca gaataatagg ttccgaaata ggcagggggc
     2101 attaactgtt tatacgggca ctgttactca aggcactgac cccgttaaaa cttattagca
     2161 gtacactcct gtatcatcaa aagccatgta tgacgcttac tggaacggta aattcagaga
     2221 ctgcgctttc cattctggct ttaatgagga tccattcgtt tgtgaatatc aaggccaatc
     2281 gtctgacctg cctcaacctc ctgtcaatgc tggcggcggc tctggtggtg gttctggtgg
     2341 cggctctgag ggtggtggct ctgagggtgg cggttctgag ggtggcggct ctgagggagg
     2401 cggttccggt ggtggctctg gttccggtga ttttgattat gaaaagatgg caaacgctaa
     2461 taagggggct atgaccgaaa atgccgatga aaacgcgcta cagtctgacg ctaaaggcaa
     2521 acttgattct gtcgctactg attacggtgc tgctatcgat ggtttcattg gtgacgtttc
     2581 cggccttgct aatggtaatg gtgctactgg tgattttgct ggctctaatt cccaaatggc
     2641 tcaagtcggt gacggtgata attcaccttt aatgaataat ttccgtcaat atttaccttc
     2701 cctccctcaa tcggttgaat gtcgcccttt tgtctttggc gctggtaaac cttacgagtt
     2761 cagtatcgac tgcgataaga tcaacctgtt ccgcggtgtc tttgcgtttc ttttatatgt
     2821 tgccaccttt atgtatgtat tttctacgtt tgctaacata ctgcgtaata aggagtctta
     2881 atgaaatttg gaaacttttt gcttacatac caacctcccc aattttccca aacagaggta
     2941 atgaaacgtt tggttaaatt aggtcgcatc tctgaggagt gtggttttga taccgtatgg
     3001 ttactggagc atcatttcac ggagtttggt ttgcttggta acccttatgt cgctgctgca
     3061 tatttacttg gcgcgactaa aaaattgaat gtaggaactg ccgctattgt tcttcccaca
     3121 gcccatccag tacgccaact tgaagatgtg aatttattgg atcaaatgtc aaaaggacga
     3181 tttcggtttg gtatttgccg agggctttac aacaaggact ttcgcgtatt cggcacagat
     3241 atgaataaca gtcgcgcctt agcggaatgc tggtacgggc tgataaagaa tggcatgaca
     3301 gagggatata tggaagctga taatgaacat atcaagttcc ataaggtaaa agtaaacccc
     3361 gcggcgtata gcagaggtgg cgcaccggtt tatgtggtgg ctgaatcagc ttcgacgact
     3421 gagtgggctg ctcaatttgg cctaccgatg atattaagtt ggattataaa tactaacgaa
     3481 aagaaagcac aacttgagct ttataatgaa gtggctcaag aatatgggca cgatattcat
     3541 aatatcgacc attgcttatc atatataaca tctgtagatc atgactcaat taaagcgaaa
     3601 gagatttgcc ggaaatttct ggggcattgg tatgattctt atgtgaatgc tacgactatt
     3661 tttgatgatt cagaccaaac aagaggttat gatttcaata aagggcagtg gcgtgacttt
     3721 gtattaaaag gacataaaga tactaatcgc cgtattgatt acagttacga aatcaatccc
     3781 gtgggaacgc cgcaggaatg tattgacata attcaaaaag acattgatgc tacaggaata
     3841 tcaaatattt gttgtggatt tgaagctaat ggaacagtag acgaaattat tgcttccatg
     3901 aagctcttcc agtctgatgt catgccattt cttaaagaaa aacaacgttc gctattatat
     3961 tatggcggtg gcggtagcgg cggtggcggt agcggcggtg gcggtagcgg cggtggcggt
     4021 agcaaatttg gattgttctt ccttaacttc atcaattcaa caactgttca agaacagagt
     4081 atagttcgca tgcaggaaat aacggagtat gttgataagt tgaattttga acagatttta
     4141 gtgtatgaaa atcatttttc agataatggt gttgtcggcg ctcctctgac tgtttctggt
     4201 tttctgctcg gtttaacaga gaaaattaaa attggttcat taaatcacat cattacaact
     4261 catcatcctg tccgcatagc ggaggaagct tgcttattgg atcagttaag tgaagggaga
     4321 tttattttag ggtttagtga ttgcgaaaaa aaagatgaaa tgcatttttt taatcgcccg
     4381 gttgaatatc aacagcaact atttgaagag tgttatgaaa tcattaacga tgctttaaca
     4441 acaggctatt gtaatccaga taacgatttt tatagcttcc ctaaaatatc tgtaaatccc
     4501 catgcttata cgccaggcgg acctcggaaa tatgtaacag caaccagtca tcatattgtt
     4561 gagtgggcgg ccaaaaaagg tattcctctc atctttaagt gggatgattc taatgatgtt
     4621 agatatgaat atgctgaaag atataaagcc gttgcggata aatatgacgt tgacctatca
     4681 gagatagacc atcagttaat gatattagtt aactataacg aagatagtaa taaagctaaa
     4741 caagagacgc gtgcatttat tagtgattat gttcttgaaa tgcaccctaa tgaaaatttc
     4801 gaaaataaac ttgaagaaat aattgcagaa aacgctgtcg gaaattatac ggagtgtata
     4861 actgcggcta agttggcaat tgaaaagtgt ggtgcgaaaa gtgtattgct gtcctttgaa
     4921 ccaatgaatg atttgatgag ccaaaaaaat gtaatcaata ttgttgatga taatattaag
     4981 aagtaccaca cggaatatac ctaaacggag ccaatgtacg caaaaaaccc cgcttcagcg
     5041 gggttttttc gccaaaaaaa accccgccct gtcaggggcg gggttttttt ttagggccct
     5101 actgactgtc cgtgtgcttc tcaaatgcct gaggccagtt tgctcaggct ctccccgtgg
     5161 aggtaataat tgacgatatg atcagtgcac ggctaactaa gcggcctgct gactttctcg
     5221 ccgatcaaaa ggcattttgc tattaaggga ttgacgaggg cgtatctgcg cagtaagatg
     5281 cgccccgcat tcggaaacct gatcatgtag atcgaatgga ctctaaatcc gttcagccgg
     5341 gttagattcc cggggtttcc gccaggatcc gtcgacctgc agatccttag cgaaagctaa
     5401 ggattttttt taagcttctg ccattcatcc gcttattatc acttattcag gcgtagcaac
     5461 caggcgttta agggcaccaa taactgcctt aaaaaaatta cgccccgccc tgccactcat
     5521 cgcagtactg ttgtaattca ttaagcattc tgccgacgtt catcggatcc cgggcccgtc
     5581 gactgcagag gcctgcatgc aagcttggcg taatcatgat ggacttaatt aacggcactc
     5641 ctcagcaaat ataatgaccc tcttgataac ccaagagggc attttttaat gcccatggcg
     5701 tttatttgcc gactaccttg gtgatctcgc ctttcacgta gtggacaaat tcttccaact
     5761 gatctgcgcg cgaggccaag cgatcttctt cttgtccaag ataagcctgt ctagcttcaa
     5821 gtatgacggg ctgatactgg gccggcaggc gctccattgc ccagtcggca gcgacatcct
     5881 tcggcgcgat tttgccggtt actgcgctgt accaaatgcg ggacaacgta agcactacat
     5941 ttcgctcatc gccagcccag tcgggcggcg agttccatag cgttaaggtt tcatttagcg
     6001 cctcaaatag atcctgttca ggaaccggat caaagagttc ctccgccgct ggacctacca
     6061 aggcaacgct atgttctctt gcttttgtca gcaagatagc cagatcaatg tcgatcgtgg
     6121 ctggctcgaa gatacctgca agaatgtcat tgcgctgcca ttctccaaat tgcagttcgc
     6181 gcttagctgg ataacgccac ggaatgatgt cgtcgtgcac aacaatggtg acttctacag
     6241 cgcggagaat ctcgctctct ccaggggaag ccgaagtttc caaaaggtcg ttgatcaaag
     6301 ctcgccgcgt tgtttcatca agccttacgg tcaccgtaac cagcaaatca atatcactgt
     6361 gtggcttcag gccgccatcc actgcggagc cgtacaaatg tacggccagc aacgtcggtt
     6421 cgagatggcg ctcgatgacg ccaactacct ctgatagttg agtcgatact tcggcgatca
     6481 ccgcttccct catactcttc ctttttcaat attattgaag catttatcag ggttattgtt
     6541 tcatgagcgt atacatattt gaatgtattt agaaaaatat taattgcgtt gcgctcactg
     6601 c
//
