LOCUS       sequence-182106-        8336 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Accessory plasmid for PACE of MjTyrRS variants; negative selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8336)
  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 8336)
  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: pDB036d
FEATURES             Location/Qualifiers
     source          1..8336
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             279..1553
                     /codon_start=1
                     /gene="III"
                     /product="pIII"
                     /label=M13 gene III
                     /note="minor coat protein"
                     /translation="MKKLLFAIPLVVPFYSHSAETVESCLAKPHTENSFTNVWKDDKTL
                     DRYANYEGCLWNATGVVVCTGDETQCYGTWVPIGLAIPENEGGGSEGGGSEGGGSEGGG
                     TKPPEYGDTPIPGYTYINPLDGTYPPGTEQNPANPNPSLEESQPLNTFMFQNNRFRNRQ
                     GALTVYTGTVTQGTDPVKTYYQYTPVSSKAMYDAYWNGKFRDCAFHSGFNEDPFVCEYQ
                     GQSSDLPQPPVNAGGGSGGGSGGGSEGGGSEGGGSEGGGSEGGGSGGGSGSGDFDYEKM
                     ANANKGAMTENADENALQSDAKGKLDSVATDYGAAIDGFIGDVSGLANGNGATGDFAGS
                     NSQMAQVGDGDNSPLMNNFRQYLPSLPQSVECRPFVFGAGKPYEFSIDCDKINLFRGVF
                     AFLLYVATFMYVFSTFANILRNKES"
     primer_bind     complement(2236..2253)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2406..2492
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2584..2611
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             complement(join(5565..6356,6357..6425))
                     /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     6188..6207
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6426..6530)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6598..6616
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             complement(6637..7587)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      7635..7857
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
ORIGIN
        1 aagaggacat ccggtcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt
       61 gctgaggaga cttagggacc ctgatgaaat tcgccacttg ttagtgtaat tcgctaactc
      121 atcctggcat gttgctgttg attcttcaat cagatcttta taaatcaaaa agataaaaaa
      181 ttggcacgca aattgtatta acagttcagc aggacaatcc tgaacgcaga aatcaagagg
      241 acaacattac ctgcaggtgc agtaaggagg aaaaaaaaat gaaaaaatta ttattcgcaa
      301 ttcctttagt tgttcctttc tattctcact ccgctgaaac tgttgaaagt tgtttagcaa
      361 aaccccatac agaaaattca tttactaacg tctggaaaga cgacaaaact ttagatcgtt
      421 acgctaacta tgagggctgt ctgtggaatg ctacaggcgt tgtagtttgt actggtgacg
      481 aaactcagtg ttacggtaca tgggttccta ttgggcttgc tatccctgaa aatgagggtg
      541 gtggctctga gggtggcggt tctgagggtg gcggttctga gggtggcggt actaaacctc
      601 ctgagtacgg tgatacacct attccgggct atacttatat caaccctctc gacggcactt
      661 atccgcctgg tactgagcaa aaccccgcta atcctaatcc ttctcttgag gagtctcagc
      721 ctcttaatac tttcatgttt cagaataata ggttccgaaa taggcagggg gcattaactg
      781 tttatacggg cactgttact caaggcactg accccgttaa aacttattac cagtacactc
      841 ctgtatcatc aaaagccatg tatgacgctt actggaacgg taaattcaga gactgcgctt
      901 tccattctgg ctttaatgag gatccattcg tttgtgaata tcaaggccaa tcgtctgacc
      961 tgcctcaacc tcctgtcaat gctggcggcg gctctggtgg tggttctggt ggcggctctg
     1021 agggtggtgg ctctgagggt ggcggttctg agggtggcgg ctctgaggga ggcggttccg
     1081 gtggtggctc tggttccggt gattttgatt atgaaaagat ggcaaacgct aataaggggg
     1141 ctatgaccga aaatgccgat gaaaacgcgc tacagtctga cgctaaaggc aaacttgatt
     1201 ctgtcgctac tgattacggt gctgctatcg atggtttcat tggtgacgtt tccggccttg
     1261 ctaatggtaa tggtgctact ggtgattttg ctggctctaa ttcccaaatg gctcaagtcg
     1321 gtgacggtga taattcacct ttaatgaata atttccgtca atatttacct tccctccctc
     1381 aatcggttga atgtcgccct tttgtctttg gcgctggtaa accttacgag ttcagtatcg
     1441 actgcgataa gatcaacctg ttccgcggtg tctttgcgtt tcttttatat gttgccacct
     1501 ttatgtatgt attttctacg tttgctaaca tactgcgtaa taaggagtct taatcttgag
     1561 taacggagcc aatgtacgca aaaaaccccg cttcagcggg gttttttcgc caaaaaaaac
     1621 cccgccctgt caggggcggg gttttttttt agggccctac tgactgtccg tgtgcttctc
     1681 aaatgcctga ggccagtttg ctcaggctct ccccgtggag gtaataattg acgatatgat
     1741 cagtgcacgg ctaactaagc ggcctgctga ctttctcgcc gatcaaaagg cattttgcta
     1801 ttaagggatt gacgagggcg tatctgcgca gtaagatgcg ccccgcattc cggcggtagt
     1861 tcagcagggc agaacggcgg actctaaatc cgcatggcag gggttcaaat cccctccgcc
     1921 ggaccaaatt cgaaaagcct gctcaacgag caggcttttt tgcatgctcg agcagctcag
     1981 ggtcgaattt gctttcgaat ttctgccatt catccgctta ttatcactta ttcaggcgta
     2041 gcaaccaggc gtttaagggc accaataact gccttaaaaa aattacgccc cgccctgcca
     2101 ctcatcgcag tactgttgta attcattaag cattctgccg acgttcatcg gatcccgggc
     2161 ccgtcgactg cagaggcctg catgcaagct tggcgtaatc agtggctgtt ttggcggatg
     2221 agagaagatt ttcagcctga tacagattaa atcagaacgc agaagcggtc tgataaaaca
     2281 gaatttgcct ggcggcagta gcgcggtggt cccacctgac cccatgccga actcagaagt
     2341 gaaactccgt agcgccgatg gtagtgtggg gtctccccat gcgagagtag ggaactgcca
     2401 gtcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt
     2461 tgtcggtgaa cgctctcctg agtaggacaa atccgccggg agcggatttg aacgttgcga
     2521 agcaacggcc cggagggtgg cgggcaggac gcccgccata aactgccagg catcaaatta
     2581 agcagaaggc catcctgacg gatggccttt ttgcgtttct acaaactcgc tgaggattct
     2641 agagcacagc aacaccacgt cgtccctatc tgctgcccta ggtctatgag tggttgctgg
     2701 ataactttac gggcatgcat aaggctcgta ggctatattc agggagacca caacggtttc
     2761 cctctacaaa taattttgtt taacttttac tagagtaagg aggaaaaatg aacacgatta
     2821 acatcgctaa gaacgacttc taggacatcg aactggctgc tatcccgttc aacactctgg
     2881 ctgaccatta cggtgagcgt ttagctcgcg aacagttggc ccttgagcat gagtcttacg
     2941 agatgggtga agcacgcttc cgcaagatgt ttgagcgtca acttaaagct ggtgaggttg
     3001 cggataacgc tgccgccaag cctctcatca ctaccctact ccctaagatg attgcacgca
     3061 tcaacgactg gtttgaggaa gtgaaagcta agcgcggcaa gcgcccgaca gccttccagt
     3121 tcctgcaaga aatcaagccg gaagccgtag cgtacatcac cattaagacc actctggctt
     3181 gcctaaccag tgctgacaat acaaccgttc aggctgtagc aagcgcaatc ggtcgggcca
     3241 ttgaggacga ggctcgcttc ggtcgtatcc gtgaccttga agctaagcac ttcaagaaaa
     3301 acgttgagga acaactcaac aagcgcgtag ggcacgtcta caagaaagca tttatgcaag
     3361 ttgtcgaggc tgacatgctc tctaagggtc tactcggtgg cgaggcgtgg tcttagtggc
     3421 ataaggaaga ctctattcat gtaggagtac gctgcatcga gatgctcatt gagtcaaccg
     3481 gaatggttag cttacaccgc caaaatgctg gcgtagtagg tcaagactct gagactatcg
     3541 aactcgcacc tgaatacgct gaggctatcg caacccgtgc aggtgcgctg gctggcatct
     3601 ctccgatgtt ccaaccttgc gtagttcctc ctaagccgtg gactggcatt actggtggtg
     3661 gctattgggc taacggtcgt cgtcctctgg cgctggtgcg tactcacagt aagaaagcac
     3721 tgatgcgcta cgaagacgtt tacatgcctg aggtgtacaa agcgattaac attgcgcaaa
     3781 acaccgcatg gaaaatcaac aagaaagtcc tagcggtcgc caacgtaatc accaagtgga
     3841 agcattgtcc ggtcgaggac atccctgcga ttgagcgtga agaactcccg atgaaaccgg
     3901 aagacatcga catgaatcct gaggctctca ccgcgtggaa acgtgctgcc gctgctgtgt
     3961 accgcaagga caaggctcgc aagtctcgcc gtatcagcct tgagttcatg cttgagcaag
     4021 ccaataagtt tgctaaccat aaggccatct ggttccctta caacatggac tggcgcggtc
     4081 gtgtttacgc tgtgtcaatg ttcaacccgc aaggtaacga tatgaccaaa ggactgctta
     4141 cgctggcgaa aggtaaacca atcggtaagg aaggttacta ctggctgaaa atccacggtg
     4201 caaactgtgc gggtgtcgat aaggttccgt tccctgagcg catcaagttc attgaggaaa
     4261 accacgagaa catcatggct tgcgctaagt ctccactgga gaacacttgg tgggctgagc
     4321 aagattctcc gttctgcttc cttgcgttct gctttgagta cgctggggta cagcaccacg
     4381 gcctgagcta taactgctcc cttccgctgg cgtttgacgg gtcttgctct ggcatccagc
     4441 acttctccgc gatgctccga gatgaggtag gtggtcgcgc ggttaacttg cttcctagtg
     4501 aaaccgttca ggacatctac gggattgttg ctaagaaagt caacgagatt ctacaagcag
     4561 acgcaatcaa tgggaccgat aacgaagtag ttaccgtgac cgatgagaac actggtgaaa
     4621 tctctgagaa agtcaagctg ggcactaagg cactggctgg tcaatggctg gcttacggtg
     4681 ttactcgcag tgtgactaag cgttcagtca tgacgctggc ttacgggtcc aaagagttcg
     4741 gcttccgtca acaagtgctg gaagatacca ttcagccagc tattgattcc ggcaagggtc
     4801 tgatgttcac tcagccgaat caggctgctg gatacatggc taagctgatt tgggaatctg
     4861 tgagcgtgac ggtggtagct gcggttgaag caatgaactg gcttaagtct gctgctaagc
     4921 tgctggctgc tgaggtcaaa gataagaaga ctggagagat tcttcgcaag cgttgcgctg
     4981 tgcattgggt aactcctgat ggtttccctg tgtggcagga atacaagaag cctattcaga
     5041 cgcgcttgaa cctgatgttc ctcggtcagt tccgcttaca gcctaccatt aacaccaaca
     5101 aagatagcga gattgatgca cacaaacagg agtctggtat cgctcctaac tttgtacaca
     5161 gccaagacgg tagccacctt cgtaagactg tagtgtgggc acacgagaag tacggaatcg
     5221 aatcttttgc actgattcac gactccttcg gtaccattcc ggctgacgct gcgaacctgt
     5281 tcaaagcagt gcgcgaaact atggttgaca catatgagtc ttgtgatgta ctggctgatt
     5341 tctacgacca gttcgctgac cagttgcacg agtctcaatt ggacaaaatg ccagcacttc
     5401 cggctaaagg taacttgaac ctccgtgaca tcttagagtc ggacttcgcg ttcgcgtaac
     5461 ttaattaacg gcactgcgaa aaaaccccgc cgaagcgggg ttttttgcga aatataatga
     5521 ccctcttgat aacccaagag ggcatttttt aatgcccatg gcgtttacca atgcttaatc
     5581 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     5641 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     5701 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     5761 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     5821 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     5881 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     5941 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     6001 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     6061 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     6121 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     6181 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     6241 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6301 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6361 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6421 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     6481 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     6541 cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat
     6601 aggcgtatca cgaggccctt aggccaaata ggccgttcag atccttccgt atttagccag
     6661 tatgttctct agtgtggttc gttgtttttg cgtgagccat gagaacgaac cattgagatc
     6721 atgcttactt tgcatgtcac tcaaaaattt tgcctcaaaa ctggtgagct gaatttttgc
     6781 agttaaagca tcgtgtagtg tttttcttag tccgttacgt aggtaggaat ctgatgtaat
     6841 ggttgttggt attttgtcac cattcatttt tatctggttg ttctcaagtt cggttacgag
     6901 atccatttgt ctatctagtt caacttggaa aatcaacgta tcagtcgggc ggcctcgctt
     6961 atcaaccacc aatttcatat tgctgtaagt gtttaaatct ttacttattg gtttcaaaac
     7021 ccattggtta agccttttaa actcatggta gttattttca agcattaaca tgaacttaaa
     7081 ttcatcaagg ctaatctcta tatttgcctt gtgagttttc ttttgtgtta gttcttttaa
     7141 taaccactca taaatcctca tagagtattt gttttcaaaa gacttaacat gttccagatt
     7201 atattttatg aattttttta actggaaaag ataaggcaat atctcttcac taaaaactaa
     7261 ttctaatttt tcgcttgaga acttggcata gtttgtccac tggaaaatct caaagccttt
     7321 aaccaaagga ttcctgattt ccacagttct cgtcatcagc tctctggttg ctttagctaa
     7381 tacaccataa gcattttccc tactgatgtt catcatctga gcgtattggt tataagtgaa
     7441 cgataccgtc cgttctttcc ttgtagggtt ttcaatcgtg gggttgagta gtgccacaca
     7501 gcataaaatt agcttggttt catgctccgt taagtcatag cgactaatcg ctagttcatt
     7561 tgctttgaaa acaactaatt cagacataca tctcaattgg tctaggtgat tttaatcact
     7621 ataccaattg agatgggcta gtcaatgata attactagtc cttttccttt gagttgtggg
     7681 tatctgtaaa ttctgctaga cctttgctgg aaaacttgta aattctgcta gaccctctgt
     7741 aaattccgct agacctttgt gtgttttttt tgtttatatt caagtggtta taatttatag
     7801 aataaagaaa gaataaaaaa agataaaaag aatagatccc agccctgtgt ataactcact
     7861 actttagtca gttccgcagt attacaaaag gatgtcgcaa acgctgtttg ctcctctaca
     7921 aaacagacct taaaacccta aaggcttaag tagcaccctc gcaagctcgg gcaaatcgct
     7981 gaatattcct tttgtctccg accatcaggc acctgagtcg ctgtcttttt cgtgacattc
     8041 agttcgctgc gctcacggct ctggcagtga atgggggtaa atggcactac aggcgccttt
     8101 tatggattca tgcaaggaaa ctacccataa tacaagaaaa gcccgtcacg ggcttctcag
     8161 ggcgttttat ggcgggtctg ctatgtggtg ctatctgact ttttgctgtt cagcagttcc
     8221 tgccctctga ttttccagtc tgaccacttc ggattatccc gtgacaggtc attcagactg
     8281 gctaatgcac ccagtaaggc agcggtatca tcaacaggct tacccgtctt actgtc
//
