LOCUS       sequence-458351-        5787 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mutant E. coli TrpRS from a derepressed lac promoter for 
            proteome-wide incorporation of tryptophan analogues in E. coli/K. 
            pneumoniae..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5787)
  AUTHORS   Loynd C, Singha Roy SJ, Canarelli SE, Bak DW, Sundaresh B, Babbitz 
            Z, van Opijnen T, Weerapana E, Chatterjee A
  TITLE     Cell-selective multiplexed bioorthogonal noncanonical amino acid 
            tagging for nascent proteomics.
  JOURNAL   Nat Chem Biol. 2025 Oct 2. doi: 10.1038/s41589-025-02039-3.
  PUBMED    41039161
REFERENCE   2  (bases 1 to 5787)
  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. 2025 Oct 2. doi: 10.1038/s41589-025-02039-3."
COMMENT     Sequence Label: pBBR1-EcTrpRS(H14)
FEATURES             Location/Qualifiers
     source          1..5787
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1023..1794
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             1795..2457
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     primer_bind     complement(4017..4033)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4017..4033)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4030..4052)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4041..4057
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4065..4071,4072..4089,4090..4095))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4110..4131
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             complement(4536..5330)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4647..4666)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5257..5276
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 ctcgggccgt ctcttgggct tgatcggcct tcttgcgcat ctcacgcgct cctgcggcgg
       61 cctgtagggc aggctcatac ccctgccgaa ccgcttttgt cagccggtcg gccacggctt
      121 ccggcgtctc aacgcgcttt gagattccca gcttttcggc caatccctgc ggtgcatagg
      181 cgcgtggctc gaccgcttgc gggctgatgg tgacgtggcc cactggtggc cgctccaggg
      241 cctcgtagaa cgcctgaatg cgcgtgtgac gtgccttgct gccctcgatg ccccgttgca
      301 gccctagatc ggccacagcg gccgcaaacg tggtctggtc gcgggtcatc tgcgctttgt
      361 tgccgatgaa ctccttggcc gacagcctgc cgtcctgcgt cagcggcacc acgaacgcgg
      421 tcatgtgcgg gctggtttcg tcacggtgga tgctggccgt cacgatgcga tccgccccgt
      481 acttgtccgc cagccacttg tgcgccttct cgaagaacgc cgcctgctgt tcttggctgg
      541 ccgacttcca ccattccggg ctggccgtca tgacgtactc gaccgccaac acagcgtcct
      601 tgcgccgctt ctctggcagc aactcgcgca gtcggcccat cgcttcatcg gtgctgctgg
      661 ccgcccagtg ctcgttctct ggcgtcctgc tggcgtcagc gttgggcgtc tcgcgctcgc
      721 ggtaggcgtg cttgagactg gccgccacgt tgcccatttt cgccagcttc ttgcatcgca
      781 tgatcgcgta tgccgccatg cctgcccctc ccttttggtg tccaaccggc tcgacggggg
      841 cagcgcaagg cggtgcctcc ggcgggccac tcaatgcttg agtatactca ctagactttg
      901 cttcgcaaag tcgtgaccgc ctacggcggc tgcggcgccc tacgggcttg ctctccgggc
      961 ttcgccctgc gcggtcgctg cgctcccttg ccagcccgtg gatatgtgga cgatggccgc
     1021 gagcggccac cggctggctc gcttcgctcg gcccgtggac aaccctgctg gacaagctga
     1081 tggacaggct gcgcctgccc acgagcttga ccacagggat tgcccaccgg ctacccagcc
     1141 ttcgaccaca tacccaccgg ctccaactgc gcggcctgcg gccttgcccc atcaattttt
     1201 ttaattttct ctggggaaaa gcctccggcc tgcggcctgc gcgcttcgct tgccggttgg
     1261 acaccaagtg gaaggcgggt caaggctcgc gcagcgaccg cgcagcggct tggccttgac
     1321 gcgcctggaa cgacccaagc ctatgcgagt gggggcagtc gaagggcgaa gcccgcccgc
     1381 ctgccccccg agcctcacgg cggcgagtgc gggggttcca agggggcagc gccaccttgg
     1441 gcaaggccga aggccgcgca gtcgatcaac aagccccgga ggggccactt tttgccggag
     1501 ggggagccgc gccgaaggcg tgggggaacc ccgcaggggt gcccttcttt gggcaccaaa
     1561 gaactagata tagggcgaaa tgcgaaagac ttaaaaatca acaacttaaa aaaggggggt
     1621 acgcaacagc tcattgcggc accccccgca atagctcatt gcgtaggtta aagaaaatct
     1681 gtaattgact gccactttta cgcaacgcat aattgttgtc gcgctgccga aaagttgcag
     1741 ctgattgcgc atggtgccgc aaccgtgcgg cacccctacc gcatggagat aagcatggcc
     1801 acgcagtcca gagaaatcgg cattcaagcc aagaacaagc ccggtcactg ggtgcaaacg
     1861 gaacgcaaag cgcatgaggc gtgggccggg cttattgcga ggaaacccac ggcggcaatg
     1921 ctgctgcatc acctcgtggc gcagatgggc caccagaacg ccgtggtggt cagccagaag
     1981 acactttcca agctcatcgg acgttctttg cggacggtcc aatacgcagt caaggacttg
     2041 gtggccgagc gctggatctc cgtcgtgaag ctcaacggcc ccggcaccgt gtcggcctac
     2101 gtggtcaatg accgcgtggc gtggggccag ccccgcgacc agttgcgcct gtcggtgttc
     2161 agtgccgccg tggtggttga tcacgacgac caggacgaat cgctgttggg gcatggcgac
     2221 ctgcgccgca tcccgaccct gtatccgggc gagcagcaac taccgaccgg ccccggcgag
     2281 gagccgccca gccagcccgg cattccgggc atggaaccag acctgccagc cttgaccgaa
     2341 acggaggaat gggaacggcg cgggcagcag cgcctgccga tgcccgatga gccgtgtttt
     2401 ctggacgatg gcgagccgtt ggagccgccg acacgggtca cgctgccgcg ccggtagcac
     2461 ttgggttgcg cagcaacccg taagtgcgct gttccagact atcggctgta gccgcctcgc
     2521 cgccctatac cttgtctgcc tccccgcgtt gcgtcgcggt gcatggagcc gggccacctc
     2581 gacctgaatg gaagccggcg gcacctcgct aacggattca ccgtttttat caggctctgg
     2641 gaggcagaat aaatgatcat atcgtcaatt attacctcca cggggagagc ctgagcaaac
     2701 tggcctcagg catttgagaa gcacacggtc acactgcttc cggtagtcaa taaaccggta
     2761 aaccagcaat agacataagc ggctatttaa cgaccctgcc ctgaaccgac gaccgggtcg
     2821 aatttgcttt cgaatttctg ccattcatcc gcttattatc acttattcag gcgtagcaac
     2881 caggcgttta agggcaccaa taactgcctt aaaaaaatta cgccccgccc tgccactcat
     2941 cgcagtacgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     3001 aacaaaatat taacgcttac ggcttcgcca caaaaccaat cgcttcgtac accgctttta
     3061 gcgtacggga agcgtgcgcg ctggcttttt ccgcgccatc tttcatcacc tgttgcagga
     3121 aggcttcatc gttgcggaaa cggtgatagc gttcctgcaa ttcagtcagc ataccggaaa
     3181 cggcatcagc cacttcacct ttcagatgac catacatctt gccttcgaac tgtttttcca
     3241 gttctgggat gctctggccc gttaccgctg aaaggatatc caacaggttg gaaacgcccg
     3301 ctttgttctg cacatcgtag cgaactaccg gcggctcgtc ggagtcagtg accgcacgtt
     3361 tgattttctt cactaccgat ttcggatctt ccagcaggcc gataacgtta ttgcgattat
     3421 cgtcagactt ggacatcttc ttggtcggct ccagcagcga cattacgcgc gcgccagatt
     3481 tcggaataaa cggctccggc accttaaaga tctcgccata cagcgcgttg aaacgctggg
     3541 caatatcgcg gctcagttcg aggtgctgtt tctggtcttc accacaagga cccagattag
     3601 tttgatacag caggatgtcc gctgccatca gcaccggata gtcaaacaga ccagcgttga
     3661 tgttctcggc ataacgcgca gatttatctt taaactgcgt catgcgactc agttcgccga
     3721 agtaggtata gcagttcagt gcccagccta actgtgcatg ttccggcacg tgggactgaa
     3781 caaaaatggt gcttttctca ggatcgatac cacaagccag atacaaggcc agcgtatcca
     3841 gcgtcgcttt acgcagcttc tgtgcatcct ggcgcacggt gatcgcgtgt tggtcaacga
     3901 tacagtaaat gcaatggtag tcatcctgca tgtttaccca ctgacgcagc gcacccatgt
     3961 agttaccaat ggtcaattca cctgagggct gtgcgccagc aaaaacgatg ggcttagtca
     4021 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     4081 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     4141 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     4201 caacgcgcgg ggagaggcgg tttgcgtatt gggcgcatgc ataaaaactg ttgtaattca
     4261 ttaagcattc tgccgacatg gaagccatca caaacggcat gatgaacctg aatcgccagc
     4321 ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca tgggggtggg cgaagaactc
     4381 cagcatgaga tccccgcgct ggaggatcat ccagccggcg tcccggaaaa cgattccgaa
     4441 gcccaacctt tcatagaagg cggcggtgga atcgaaatct cgtgatggca ggttgggcgt
     4501 cgcttggtcg gtcatttcga accccagagt cccgctcaga agaactcgtc aagaaggcga
     4561 tagaaggcga tgcgctgcga atcgggagcg gcgataccgt aaagcacgag gaagcggtca
     4621 gcccattcgc cgccaagctc ttcagcaata tcacgggtag ccaacgctat gtcctgatag
     4681 cggtccgcca cacccagccg gccacagtcg atgaatccag aaaagcggcc attttccacc
     4741 atgatattcg gcaagcaggc atcgccatgg gtcacgacga gatcctcgcc gtcgggcatg
     4801 cgcgccttga gcctggcgaa cagttcggct ggcgcgagcc cctgatgctc ttcgtccaga
     4861 tcatcctgat cgacaagacc ggcttccatc cgagtacgtg ctcgctcgat gcgatgtttc
     4921 gcttggtggt cgaatgggca ggtagccgga tcaagcgtat gcagccgccg cattgcatca
     4981 gccatgatgg atactttctc ggcaggagca aggtgagatg acaggagatc ctgccccggc
     5041 acttcgccca atagcagcca gtcccttccc gcttcagtga caacgtcgag cacagctgcg
     5101 caaggaacgc ccgtcgtggc cagccacgat agccgcgctg cctcgtcctg cagttcattc
     5161 agggcaccgg acaggtcggt cttgacaaaa agaaccgggc gcccctgcgc tgacagccgg
     5221 aacacggcgg catcagagca gccgattgtc tgttgtgccc agtcatagcc gaatagcctc
     5281 tccacccaag cggccggaga acctgcgtgc aatccatctt gttcaatcat gcgaaacgat
     5341 cctcatcctg tctcttgatc agatcttgat cccctgcgcc atcagatcct tggcggcaag
     5401 aaagccatcc agtttacttt gcagggcttc ccaaccttac cagagggcgc cccagctggc
     5461 aattccggtt cgcttgctgt ccataaaacc gcccagtcta gctatcgcca tgtaagccca
     5521 ctgcaagcta cctgctttct ctttgcgctt gcgttttccc ttgtccagat agcccagtag
     5581 ctgacattca tcccaggtgg cacttttcgg ggaaatgtgc gcgcccgcgt tcctgctggc
     5641 gctgggcctg tttctggcgc tggacttccc gctgttccgt cagcagcttt tcgcccacgg
     5701 ccttgatgat cgcggcggcc ttggcctgca tatcccgatt caacggcccc agggcgtcca
     5761 gaacgggctt caggcgctcc cgaaggt
//
