LOCUS       sequence-182099-        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"
     primer_bind     489..508
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     742..759
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    774..914
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(840..859)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1000..1022
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1016..1207)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             2856..3935
                     /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             3999..4979
                     /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(5677..6468)
                     /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(6469..6540)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      join(6601,1..588)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
       61 gcggtggttt gtttgccgga tcaagagcta ccacctcttt ttccgaaggt aactggcttc
      121 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
      181 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
      241 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
      301 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
      361 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
      421 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
      481 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
      541 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
      601 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
      661 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
      721 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
      781 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatggtg cactctcagt
      841 acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact
      901 gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
      961 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     1021 ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt
     1081 cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca
     1141 gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt
     1201 tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga
     1261 aacgagagag gatgctcacg atacgggtta ctgatgatga acaaagagga catccggtca
     1321 aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttgctgagg agacttaggg
     1381 accctgatga aattcgccac ttgttagtgt aattcgctaa ctcatcctgg catgttgctg
     1441 ttgattcttc aatcagatct ttataaatca aaaagataaa aaattggcac gcaaattgta
     1501 ttaacagttc agcaggacaa tcctgaacgc agaaatcaag aggacaacat tacctgcagg
     1561 tgcagtaagg aggaaaaaaa aatgaaaaaa ttattattcg caattccttt agttgttcct
     1621 ttctattctc actccgctga aactgttgaa agttgtttag caaaatagca tacagaaaat
     1681 tcatttacta acgtctggaa agacgacaaa actttagatc gttacgctaa ctatgagggc
     1741 tgtctgtgga atgctacagg cgttgtagtt tgtactggtg acgaaactca gtgttacggt
     1801 acatgggttc ctattgggct tgctatctag gaaaatgagg gtggtggctc tgagggtggc
     1861 ggttctgagg gtggcggttc tgagggtggc ggtactaaac ctcctgagta cggtgataca
     1921 cctattccgg gctatactta tatcaaccct ctcgacggca cttatccgcc tggtactgag
     1981 caaaaccccg ctaatcctaa tccttctctt gaggagtctc agcctcttaa tactttcatg
     2041 tttcagaata ataggttccg aaataggcag ggggcattaa ctgtttatac gggcactgtt
     2101 actcaaggca ctgaccccgt taaaacttat tagcagtaca ctcctgtatc atcaaaagcc
     2161 atgtatgacg cttactggaa cggtaaattc agagactgcg ctttccattc tggctttaat
     2221 gaggatccat tcgtttgtga atatcaaggc caatcgtctg acctgcctca acctcctgtc
     2281 aatgctggcg gcggctctgg tggtggttct ggtggcggct ctgagggtgg tggctctgag
     2341 ggtggcggtt ctgagggtgg cggctctgag ggaggcggtt ccggtggtgg ctctggttcc
     2401 ggtgattttg attatgaaaa gatggcaaac gctaataagg gggctatgac cgaaaatgcc
     2461 gatgaaaacg cgctacagtc tgacgctaaa ggcaaacttg attctgtcgc tactgattac
     2521 ggtgctgcta tcgatggttt cattggtgac gtttccggcc ttgctaatgg taatggtgct
     2581 actggtgatt ttgctggctc taattcccaa atggctcaag tcggtgacgg tgataattca
     2641 cctttaatga ataatttccg tcaatattta ccttccctcc ctcaatcggt tgaatgtcgc
     2701 ccttttgtct ttggcgctgg taaaccttac gagttcagta tcgactgcga taagatcaac
     2761 ctgttccgcg gtgtctttgc gtttctttta tatgttgcca cctttatgta tgtattttct
     2821 acgtttgcta acatactgcg taataaggag tcttaatgaa atttggaaac tttttgctta
     2881 cataccaacc tccccaattt tcccaaacag aggtaatgaa acgtttggtt aaattaggtc
     2941 gcatctctga ggagtgtggt tttgataccg tatggttact ggagcatcat ttcacggagt
     3001 ttggtttgct tggtaaccct tatgtcgctg ctgcatattt acttggcgcg actaaaaaat
     3061 tgaatgtagg aactgccgct attgttcttc ccacagccca tccagtacgc caacttgaag
     3121 atgtgaattt attggatcaa atgtcaaaag gacgatttcg gtttggtatt tgccgagggc
     3181 tttacaacaa ggactttcgc gtattcggca cagatatgaa taacagtcgc gccttagcgg
     3241 aatgctggta cgggctgata aagaatggca tgacagaggg atatatggaa gctgataatg
     3301 aacatatcaa gttccataag gtaaaagtaa accccgcggc gtatagcaga ggtggcgcac
     3361 cggtttatgt ggtggctgaa tcagcttcga cgactgagtg ggctgctcaa tttggcctac
     3421 cgatgatatt aagttggatt ataaatacta acgaaaagaa agcacaactt gagctttata
     3481 atgaagtggc tcaagaatat gggcacgata ttcataatat cgaccattgc ttatcatata
     3541 taacatctgt agatcatgac tcaattaaag cgaaagagat ttgccggaaa tttctggggc
     3601 attggtatga ttcttatgtg aatgctacga ctatttttga tgattcagac caaacaagag
     3661 gttatgattt caataaaggg cagtggcgtg actttgtatt aaaaggacat aaagatacta
     3721 atcgccgtat tgattacagt tacgaaatca atcccgtggg aacgccgcag gaatgtattg
     3781 acataattca aaaagacatt gatgctacag gaatatcaaa tatttgttgt ggatttgaag
     3841 ctaatggaac agtagacgaa attattgctt ccatgaagct cttccagtct gatgtcatgc
     3901 catttcttaa agaaaaacaa cgttcgctat tatattatgg cggtggcggt agcggcggtg
     3961 gcggtagcgg cggtggcggt agcggcggtg gcggtagcaa atttggattg ttcttcctta
     4021 acttcatcaa ttcaacaact gttcaagaac agagtatagt tcgcatgcag gaaataacgg
     4081 agtatgttga taagttgaat tttgaacaga ttttagtgta tgaaaatcat ttttcagata
     4141 atggtgttgt cggcgctcct ctgactgttt ctggttttct gctcggttta acagagaaaa
     4201 ttaaaattgg ttcattaaat cacatcatta caactcatca tcctgtccgc atagcggagg
     4261 aagcttgctt attggatcag ttaagtgaag ggagatttat tttagggttt agtgattgcg
     4321 aaaaaaaaga tgaaatgcat ttttttaatc gcccggttga atatcaacag caactatttg
     4381 aagagtgtta tgaaatcatt aacgatgctt taacaacagg ctattgtaat ccagataacg
     4441 atttttatag cttccctaaa atatctgtaa atccccatgc ttatacgcca ggcggacctc
     4501 ggaaatatgt aacagcaacc agtcatcata ttgttgagtg ggcggccaaa aaaggtattc
     4561 ctctcatctt taagtgggat gattctaatg atgttagata tgaatatgct gaaagatata
     4621 aagccgttgc ggataaatat gacgttgacc tatcagagat agaccatcag ttaatgatat
     4681 tagttaacta taacgaagat agtaataaag ctaaacaaga gacgcgtgca tttattagtg
     4741 attatgttct tgaaatgcac cctaatgaaa atttcgaaaa taaacttgaa gaaataattg
     4801 cagaaaacgc tgtcggaaat tatacggagt gtataactgc ggctaagttg gcaattgaaa
     4861 agtgtggtgc gaaaagtgta ttgctgtcct ttgaaccaat gaatgatttg atgagccaaa
     4921 aaaatgtaat caatattgtt gatgataata ttaagaagta ccacacggaa tatacctaaa
     4981 cggagccaat gtacgcaaaa aaccccgctt cagcggggtt ttttcgccaa aaaaaacccc
     5041 gccctgtcag gggcggggtt tttttttagg gccctactga ctgtccgtgt gcttctcaaa
     5101 tgcctgaggc cagtttgctc aggctctccc cgtggaggta ataattgacg atatgatcag
     5161 tgcacggcta actaagcggc ctgctgactt tctcgccgat caaaaggcat tttgctatta
     5221 agggattgac gagggcgtat ctgcgcagta agatgcgccc cgcattcgga aacctgatca
     5281 tgtagatcga atggactcta aatccgttca gccgggttag attcccgggg tttccgccag
     5341 gatccgtcga cctgcagatc cttagcgaaa gctaaggatt ttttttaagc ttctgccatt
     5401 catccgctta ttatcactta ttcaggcgta gcaaccaggc gtttaagggc accaataact
     5461 gccttaaaaa aattacgccc cgccctgcca ctcatcgcag tactgttgta attcattaag
     5521 cattctgccg acgttcatcg gatcccgggc ccgtcgactg cagaggcctg catgcaagct
     5581 tggcgtaatc atgatggact taattaacgg cactcctcag caaatataat gaccctcttg
     5641 ataacccaag agggcatttt ttaatgccca tggcgtttat ttgccgacta ccttggtgat
     5701 ctcgcctttc acgtagtgga caaattcttc caactgatct gcgcgcgagg ccaagcgatc
     5761 ttcttcttgt ccaagataag cctgtctagc ttcaagtatg acgggctgat actgggccgg
     5821 caggcgctcc attgcccagt cggcagcgac atccttcggc gcgattttgc cggttactgc
     5881 gctgtaccaa atgcgggaca acgtaagcac tacatttcgc tcatcgccag cccagtcggg
     5941 cggcgagttc catagcgtta aggtttcatt tagcgcctca aatagatcct gttcaggaac
     6001 cggatcaaag agttcctccg ccgctggacc taccaaggca acgctatgtt ctcttgcttt
     6061 tgtcagcaag atagccagat caatgtcgat cgtggctggc tcgaagatac ctgcaagaat
     6121 gtcattgcgc tgccattctc caaattgcag ttcgcgctta gctggataac gccacggaat
     6181 gatgtcgtcg tgcacaacaa tggtgacttc tacagcgcgg agaatctcgc tctctccagg
     6241 ggaagccgaa gtttccaaaa ggtcgttgat caaagctcgc cgcgttgttt catcaagcct
     6301 tacggtcacc gtaaccagca aatcaatatc actgtgtggc ttcaggccgc catccactgc
     6361 ggagccgtac aaatgtacgg ccagcaacgt cggttcgaga tggcgctcga tgacgccaac
     6421 tacctctgat agttgagtcg atacttcggc gatcaccgct tccctcatac tcttcctttt
     6481 tcaatattat tgaagcattt atcagggtta ttgtttcatg agcgtataca tatttgaatg
     6541 tatttagaaa aatattaatt gcgttgcgct cactgcccgc tttccaggcc aaataggccg
     6601 t
//
