LOCUS       sequence-341130-        6988 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for incorporating the non-canonical amino acid 
            3-nitroTyrosine with the third generation Mj 3NY (A7) synthetase and
            cognate amber suppressing tRNA in Ecoli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6988)
  AUTHORS   Beyer JN, Hosseinzadeh P, Gottfried-Lee I, Van Fossen EM, Zhu P, 
            Bednar RM, Karplus PA, Mehl RA, Cooley RB
  TITLE     Overcoming Near-Cognate Suppression in a Release Factor 1-Deficient 
            Host with an Improved Nitro-Tyrosine tRNA Synthetase.
  JOURNAL   J Mol Biol. 2020 Jul 24;432(16):4690-4704. doi: 
            10.1016/j.jmb.2020.06.014. Epub 2020 Jun 19.
  PUBMED    32569745
REFERENCE   2  (bases 1 to 6988)
  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"; doi: 
            "10.1016/j.jmb.2020.06.014"; journalName: "J Mol Biol"; date: 
            "2020-07-24- 24"; volume: "432"; issue: "16"; pages: "4690-4704"
COMMENT     Sequence Label: pDule2-3-nitroTyrosine (A7)
FEATURES             Location/Qualifiers
     source          1..6988
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1168..1197
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1213..1230
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1286..1303)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1456..1542
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(1749..1766)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1883..2428)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(1963..1982)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2579..2670
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2671..3462
                     /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"
     primer_bind     complement(3638..3657)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     4783..4805
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4798..4814
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5798..5820
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5976..6854)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 ggacagcggt gcggactgtt gtaactcaga ataagaaatg aggccgctca tggcgttctg
       61 ttgcccgtct caccggtgaa aagaaaaaca accctggcgc cgcttctttg agcgaacgat
      121 caaaaataag tggcgcccca tcaaaaaaat attctcaaca taaaaaactt tgtgtaatac
      181 ttgtaacgct gagtttacgc tttgaggaat cccccatgga cgaatttgaa atgataaaga
      241 gaaacacatc tgaaattatc agcgaggaag agttaagaga ggttttaaaa aaagatgaaa
      301 aatctgctca tataggtttt gaaccaagtg gtaaaataca tttagggcat tatctccaaa
      361 taaaaaagat gattgattta caaaatgctg gatttgatat aattatactt ttggctgatt
      421 taacggccta tttaaaccag aaaggagagt tggatgagat tagaaaaata ggagattata
      481 acaaaaaagt ttttgaagca atggggttaa aggcaaaata tgtttatgga agtgaattcc
      541 agcttgataa ggattataca ctgaatgtct atagattggc tttaaaaact accttaaaaa
      601 gagcaagaag gagtatggaa cttatagcaa gagaggatga aaatccaaag gttgctgaag
      661 ttatctatcc aataatgcag gttaatcatg ctcattatcg tggcgttgat gttgcagttg
      721 gagggatgga gcagagaaaa atacacatgt tagcaaggga gcttttacca aaaaaggttg
      781 tttgtattca caaccctgtc ttaacgggtt tggatggaga aggaaagatg agttcttcaa
      841 aagggaattt tatagctgtt gatgactctc cagaagagat tagggctaag ataaagaaag
      901 catactgccc agctggagtt gttgaaggaa atccaataat ggagatagct aaatacttcc
      961 ttgaatatcc tttaaccata aaaaggccag aaaaatttgg tggagatttg acagttaata
     1021 gctatgagga gttagagagt ttatttaaaa ataaggaatt gcatccaatg gatttaaaaa
     1081 atgctgtagc tgaagaactt ataaagattt tagagccaat tagaaagaga ttatgaggta
     1141 ccatatggga attcgaagct tgggcccgaa caaaaactca tctcagaaga ggatctgaat
     1201 agcgccgtcg accatcatca tcatcatcat tgagtttaaa cggtctccag cttggctgtt
     1261 ttggcggatg agagaagatt ttcagcctga tacagattaa atcagaacgc agaagcggtc
     1321 tgataaaaca gaatttgcct ggcggcagta gcgcggtggt cccacctgac cccatgccga
     1381 actcagaagt gaaacgccgt agcgccgatg gtagtgtggg gtctccccat gcgagagtag
     1441 ggaactgcca ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt
     1501 atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa atccgccggg agctgtccct
     1561 cctgttcagc tactgacggg gtggtgcgta acggcaaaag caccgccgga catcagcgct
     1621 agcggagtgt atactggctt actatgttgg cactgatgag ggtgtcagtg aagtgcttca
     1681 tgtggcagga gaaaaaaggc tgcaccggtg cgtcagcaga atatgtgata caggatatat
     1741 tccgcttcct cgctcactga ctcgctacgc tcggtcgttc gactgcggcg agcggaaatg
     1801 gcttacgaac ggggcggaga tttcctggaa gatgccagga agatacttaa cagggaagtg
     1861 agagggccgc ggcaaagccg tttttccata ggctccgccc ccctgacaag catcacgaaa
     1921 tctgacgctc aaatcagtgg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     1981 cccctggcgg ctccctcgtg cgctctcctg ttcctgcctt tcggtttacc ggtgtcattc
     2041 cgctgttatg gccgcgtttg tctcattcca cgcctgacac tcagttccgg gtaggcagtt
     2101 cgctccaagc tggactgtat gcacgaaccc cccgttcagt ccgaccgctg cgccttatcc
     2161 ggtaactatc gtcttgagtc caacccggaa agacatgcaa aagcaccact ggcagcagcc
     2221 actggtaatt gatttagagg agttagtctt gaagtcatgc gccggttaag gctaaactga
     2281 aaggacaagt tttggtgact gcgctcctcc aagccagtta cctcggttca aagagttggt
     2341 agctcagaga accttcgaaa aaccgccctg caaggcggtt ttttcgtttt cagagcaaga
     2401 gattacgcgc agaccaaaac gatctcaaga agatcatctt attaatcaga taaaatattt
     2461 ctagatttca gtgcaattta tctcttcaaa tgtagcacct gaagtcagcc ccatacgata
     2521 taagttgtaa ttctcatgtt tgacagctta tcatcgatgc gaccgagtga gctagctatt
     2581 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     2641 atgcttcaat aatattgaaa aaggaagagt atgagggaag cggtgatcgc cgaagtatcg
     2701 actcaactat cagaggtagt tggcgtcatc gagcgccatc tcgaaccgac gttgctggcc
     2761 gtacatttgt acggctccgc agtggatggc ggcctgaagc cacacagtga tattgatttg
     2821 ctggttacgg tgaccgtaag gcttgatgaa acaacgcggc gagctttgat caacgacctt
     2881 ttggaaactt cggcttcccc tggagagagc gagattctcc gcgctgtaga agtcaccatt
     2941 gttgtgcacg acgacatcat tccgtggcgt tatccagcta agcgcgaact gcaatttgga
     3001 gaatggcagc gcaatgacat tcttgcaggt atcttcgagc cagccacgat cgacattgat
     3061 ctggctatct tgctgacaaa agcaagagaa catagcgttg ccttggtagg tccagcggcg
     3121 gaggaactct ttgatccggt tcctgaacag gatctatttg aggcgctaaa tgaaacctta
     3181 acgctatgga actcgccgcc cgactgggct ggcgatgagc gaaatgtagt gcttacgttg
     3241 tcccgcattt ggtacagcgc agtaaccggc aaaatcgcgc cgaaggatgt cgctgccgac
     3301 tgggcaatgg agcgcctgcc ggcccagtat cagcccgtca tacttgaagc tagacaggct
     3361 tatcttggac aagaagaaga tcgcttggcc tcgcgcgcag atcagttgga agaatttgtc
     3421 cactacgtga aaggcgagat caccaaggta gtcggcaaat aatgtctaac aattcgttca
     3481 agccgagggg ccgcaagatc cggccacgat gacccggtcg tcggttcagg gcagggtcgt
     3541 taaatagccg cttatgtcta ttgctggttt accgggatcc tctacgccgg acgcatcgtg
     3601 gccggcatca ccggcgccac aggtgcggtt gctggcgcct atatcgccga catcaccgat
     3661 ggggaagatc gggctcgcca cttcgggctc atgagcgctt gtttcggcgt gggtatggtg
     3721 gcaggccccg tggccggggg actgttgggc gccatctcct tgcatgcacc attccttgcg
     3781 gcggcggtgc tcaacggcct caacctacta ctgggctgct tcctaatgca ggagtcgcat
     3841 aagggagagc gtcgaccgat gcccttgaga gccttcaacc cagtcagctc cttccggtgg
     3901 gcgcggggca tgactatcgt cgccgcactt atgactgtct tctttatcat gcaactcgta
     3961 ggacaggtgc cggcagcgct ctgggtcatt ttcggcgagg accgctttcg ctggagcgcg
     4021 acgatgatcg gcctgtcgct tgcggtattc ggaatcttgc acgccctcgc tcaagccttc
     4081 gtcactggtc ccgccaccaa acgtttcggc gagaagcagg ccattatcgc cggcatggcg
     4141 gccgacgcgc tgggctacgt cttgctggcg ttcgcgacgc gaggctggat ggccttcccc
     4201 attatgattc ttctcgcttc cggcggcatc gggatgcccg cgttgcaggc catgctgtcc
     4261 aggcaggtag atgacgacca tcagggacag cttcaaggat cgctcgcggc tcttaccagc
     4321 ctaacttcga tcattggacc gctgatcgtc acggcgattt atgccgcctc ggcgagcaca
     4381 tggaacgggt tggcatggat tgtaggcgcc gccctatacc ttgtctgcct ccccgcgttg
     4441 cgtcgcggtg catggagccg ggccacctcg acctgaatgg aagccggcgg cacctcgcta
     4501 acggattcac cactccaaga attggagcca atcaattctt gcggagaact gtgaatgcgc
     4561 aaaccaaccc ttggcagaac atatccatcg cgtccgccat ctccagcagc cgcacgcggc
     4621 gcatctcggg ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc
     4681 tactactggg ctgcttccta atgcaggagt cgcataaggg agagcgtctg gcgaaagggg
     4741 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
     4801 aaacgacggc cagtgccaag cttaaaaaaa atccttagct ttcgctaagg atctgcagtg
     4861 gtccggcggg ccggatttga accagcgcca tgcggattta gagtccgccg ttctgccctg
     4921 ctgaactacc gccgggaatt cagcgttaca agtattacac aaagtttttt atgttgagaa
     4981 tatttttttg atggggcgcc acttattttt gatcgttcgc tcaaagaagc ggcgccaggg
     5041 ttgtttttct tttcaccagt gagacgggca acagaacgcc atgagcggcc tcatttctta
     5101 ttctgagtta caacagtccg caccgctgcc ggtagctcct tccggtgggc gcggggcatg
     5161 actatcgtcg ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg
     5221 gcagcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg
     5281 ccctgcacca ttatgttccg gatctgcatc gcaggatgct gctggctacc ctgtggaaca
     5341 cctacatctg tattaacgaa gcgctaaccg tttttatcat gctctgggag gcagaataaa
     5401 tgatcatatc gtcaattatt acctccacgg ggagagcctg agcaaactgg cctcaggcat
     5461 ttgagaagca cacggtcaca ctgcttccgg tagtcaataa accggtaaac cagcaataga
     5521 cataagcggc tatttaacga ccctgccctg aaccgacgac cgggtcgaat ttgctttcga
     5581 atttctgcca ttcatccgct tattatcact tattcaggcg tagcaaccag gcgtttaagg
     5641 gcaccaataa ctgccttaaa aaaattacgc cccgccctgc cactcatcgc agttgactgg
     5701 gtcatggctg cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     5761 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     5821 ttttcaccgt catcaccgaa acgcgcgagg cagcagatca attcgcgcgc gaaggcgaag
     5881 cggcatgcat aatgtgcctg tcaaatggac gaagcaggga ttctgcaaac cctatgctac
     5941 tccgtcaagc cgtcaattgt ctgattcgtt accaattatg acaacttgac ggctacatca
     6001 ttcacttttt cttcacaacc ggcacggaac tcgctcgggc tggccccggt gcatttttta
     6061 aatacccgcg agaaatagag ttgatcgtca aaaccaacat tgcgaccgac ggtggcgata
     6121 ggcatccggg tggtgctcaa aagcagcttc gcctggctga tacgttggtc ctcgcgccag
     6181 cttaagacgc taatccctaa ctgctggcgg aaaagatgtg acagacgcga cggcgacaag
     6241 caaacatgct gtgcgacgct ggcgatatca aaattgctgt ctgccaggtg atcgctgatg
     6301 tactgacaag cctcgcgtac ccgattatcc atcggtggat ggagcgactc gttaatcgct
     6361 tccatgcgcc gcagtaacaa ttgctcaagc agatttatcg ccagcagctc cgaatagcgc
     6421 ccttcccctt gcccggcgtt aatgatttgc ccaaacaggt cgctgaaatg cggctggtgc
     6481 gcttcatccg ggcgaaagaa ccccgtattg gcaaatattg acggccagtt aagccattca
     6541 tgccagtagg cgcgcggacg aaagtaaacc cactggtgat accattcgcg agcctccgga
     6601 tgacgaccgt agtgatgaat ctctcctggc gggaacagca aaatatcacc cggtcggcaa
     6661 acaaattctc gtccctgatt tttcaccacc ccctgaccgc gaatggtgag attgagaata
     6721 taacctttca ttcccagcgg tcggtcgata aaaaaatcga gataaccgtt ggcctcaatc
     6781 ggcgttaaac ccgccaccag atgggcatta aacgagtatc ccggcagcag gggatcattt
     6841 tgcgcttcag ccatactttt catactcccg ccattcagag aagaaaccaa ttgtccatat
     6901 tgcatcagac attgccgtca ctgcgtcttt tactggctct tctcgctaac caaaccggta
     6961 acccgggacc aagtttactc atatatac
//
