LOCUS       sequence-494133-        7273 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression plasmid encoding the evolved MjTyrRS variant Iodo.5RS for
            amber suppression with p-iodophenylalanine.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7273)
  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 7273)
  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: pDB070.iodo.5
FEATURES             Location/Qualifiers
     source          1..7273
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(1..546)
                     /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.
                     "
     CDS             complement(694..1317)
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     1225..1244
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    1425..1443
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        1444..1498
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    1449..1467
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    1479..1497
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     complement(2469..2486)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2639..2685
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(3395..4054)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        complement(4055..4157)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     rep_origin      complement(4683..5228)
                     /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.
                     "
     CDS             complement(5986..6645)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        complement(6646..6748)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
ORIGIN
        1 tttccatagg ctccgccccc ctgacaagca tcacgaaatc tgacgctcaa atcagtggtg
       61 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggcggct ccctcgtgcg
      121 ctctcctgtt cctgcctttc ggtttaccgg tgtcattccg ctgttatggc cgcgtttgtc
      181 tcattccacg cctgacactc agttccgggt aggcagttcg ctccaagctg gactgtatgc
      241 acgaaccccc cgttcagtcc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
      301 acccggaaag acatgcaaaa gcaccactgg cagcagccac tggtaattga tttagaggag
      361 ttagtcttga agtcatgcgc cggttaaggc taaactgaaa ggacaagttt tggtgactgc
      421 gctcctccaa gccagttacc tcggttcaaa gagttggtag ctcagagaac cttcgaaaaa
      481 ccgccctgca aggcggtttt ttcgttttca gagcaagaga ttacgcgcag accaaaacga
      541 tctcaagaag atcatcttat taatcagata aaatatttct agatttcagt gcaatttatc
      601 tcttcaaatg tagcacctga agtcagcccc atacgatata agttgtaatt ctcatgtttg
      661 acagcttatc atcgataaag ctatctagaa agcttaagac ccactttcac atttaagttg
      721 tttttctaat ccgcatatga tcaattcaag gccgaataag aaggctggct ctgcaccttg
      781 gtgatcaaat aattcgatag cttgtcgtaa taatggcggc atactatcag tagtaggtgt
      841 ttccctttct tctttagcga cttgatgctc ttgatcttcc aatacgcaac ctaaagtaaa
      901 atgccccact gcgctgagtg catataatgc attctctagt gaaaaacctt gttggcataa
      961 aaaggctaat tgattttcga gagtttcata ctgtttttct gtaggccgtg tacctaaatg
     1021 tacttttgct ccatcgcgat gacttagtaa agcacatcta aaacttttag cgttattacg
     1081 taaaaaatct tgccagcttt ccccttctaa agggcaaaag tgagtatggt gcctatctaa
     1141 catctcaatg gctaaggcgt cgagcaaagc ccgcttattt tttacatgcc aatacaatgt
     1201 aggctgctct acacctagct tctgggcgag tttacgggtt gttaaacctt cgattccgac
     1261 ctcattaagc agctctaatg cgctgttaat cactttactt ttatctaatc tagacatatg
     1321 aattcggggc tttatgaatc tattatacag aaaaattttc ctgaaagcaa ataattaatt
     1381 gcgttgcgca ttctagagca cagctccatc gaatggccag atgatcccta tcagtgatag
     1441 agattgacac tctatcattg atagagttat tttaccactc cctatcagtg atagagaaag
     1501 acctatggac gaatttgaaa tgataaagag aaacacatct gaaattatca gcgaggaaga
     1561 gttaagagag gttttaaaaa aagatgaaaa atctgctctg ataggttttg aaccaagtgg
     1621 taaaatacat ttagggcatt atctccaaat aaaaaagatg attgatttac aaaatgctgg
     1681 atttgatata attatattgt tggctgattt ccacgcctat ttaaaccaga aaggagagtt
     1741 ggatgagatt agaaaaatag gagattataa caaaaaagtt tttgaagcaa tggggttaaa
     1801 ggcaaaatat gtttatggaa gttcgttcca gcttgataag gattatacac tgaatgtcta
     1861 tagattggct ttaaaaacta ccttaaaaag agcaagaagg agtatggaac ttatagcaag
     1921 agaggatgaa aatccaaagg ttgctgaagt tatctatcca ataatgcagg ttaatcctct
     1981 taattatgag ggcgttgatg ttgcagttgg agggatggag cagagaaaaa tacacatgtt
     2041 agcaagggag cttttaccaa aaaaggttgt ttgtattcac aaccctgtct taacgggttt
     2101 ggatggagaa ggaaagatga gttcttcaaa agggaatttt atagctgttg atgactctcc
     2161 agaagagatt agggctaaga taaagaaagc atactgccca gctggaattg ttgaaggaaa
     2221 tccaataatg gagatagcta aatacttcct tgaatatcct ttaaccataa aaaggccaga
     2281 aaaatttggt ggagatttga cagttaatag ctatgaggag ttagagagtt tatttaaaaa
     2341 taaggaattg catccaatgc gcttaaaaaa tgctgtagct gaagaactta taaagatttt
     2401 agagccaatt agaaagagat tataagtctc cagcttggct gttttggcgg atgagagaag
     2461 attttcagcc tgatacagat taaatcagaa cgcagaagcg gtctgataaa acagaatttg
     2521 cctggcggca gtagcgcggt ggtcccacct gaccccatgc cgaactcaga agtgaaacgc
     2581 cgtagcgccg atggtagtgt ggggtctccc catgcgagag tagggaactg ccaggcatca
     2641 aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtgagc
     2701 tcccggtcat caatcatccc cataatcctt gttagattat caaagtttca aacgctaaat
     2761 tgcctgatgc gctacgctta tcaggcctac atgatctctg caatatattg agtttgcgtg
     2821 cttttgtagg ccggataagg cgttcacgcc gcatccggca agaaacagca aacaatccaa
     2881 aacgccgcgt tcagcggcgt tttttctgct tttcttcgcg aattaattcc gcttcgcaac
     2941 atgtgagcac cggtttattg actaccggaa gcagtgtgac cgtgtgcttc tcaaatgcct
     3001 gaggccagtt tgctcaggct ctccccgtgg aggtaataat tgacgatatg atcagtgcac
     3061 ggctaactaa gcggcctgct gactttctcg ccgatcaaaa ggcattttgc tattaaggga
     3121 ttgacgaggg cgtatctgcg cagtaagatg cgccccgcat tccggcggta gttcagcagg
     3181 gcagaacggc ggactctaaa tccgcatggc aggggttcaa atcccctccg ccggaccaaa
     3241 ttcgaaaagc ctgctcaacg agcaggcttt tttgcatgct cgagcagctc agggtcgaat
     3301 ttgctttcga atttctgcca ttcatccgct tattatcact tattcaggcg tagcaaccag
     3361 gcgtttaagg gcaccaataa ctgccttaaa aaaattacgc cccgccctgc cactcatcgc
     3421 agtactgttg taattcatta agcattctgc cgacatggaa gccatcacaa acggcatgat
     3481 gaacctgaat cgccagcggc atcagcacct tgtcgccttg cgtataatat ttgcccatgg
     3541 tgaaaacggg ggcgaagaag ttgtccatat tggccacgtt taaatcaaaa ctggtgaaac
     3601 tcacccaggg attggctgag acgaaaaaca tattctcaat aaacccttta gggaaatagg
     3661 ccaggttttc accgtaacac gccacatctt gcgaatatat gtgtagaaac tgccggaaat
     3721 cgtcgtggta ttcactccag agcgatgaaa acgtttcagt ttgctcatgg aaaacggtgt
     3781 aacaagggtg aacactatcc catatcacca gctcaccgtc tttcattgcc atacggaatt
     3841 ccggatgagc attcatcagg cgggcaagaa tgtgaataaa ggccggataa aacttgtgct
     3901 tatttttctt tacggtcttt aaaaaggccg taatatccag ctgaacggtc tggttatagg
     3961 tacattgagc aactgactga aatgcctcaa aatgttcttt acgatgccat tgggatatat
     4021 caacggtggt atatccagtg atttttttct ccattttagc ttccttagct cctgaaaatc
     4081 tcgataactc aaaaaatacg cccggtagtg atcttatttc attatggtga aagttggaac
     4141 ctcttacgtg ccgatcaacg tctcattttc gccaaaagtt ggcccagggc ttcccggtat
     4201 caacagggac accaggattt atttattctg cgaagtgatc ttccgtcaca ggtatttatt
     4261 cggcgcaaag tgcgtcgggt gatgctgcca acttactgat ttagtgtatg atggtgtttt
     4321 tgaggtgctc cagtggcttc tgtttctatc agctgtccct cctgttcagc tactgacggg
     4381 gtggtgcgta acggcaaaag caccgccgga catcagcgct agcggagtgt atactggctt
     4441 actatgttgg cactgatgag ggtgtcagtg aagtgcttca tgtggcagga gaaaaaaggc
     4501 tgcaccggtg cgtcagcaga atatgtgata caggatatat tccgcttcct cgctcactga
     4561 ctcgctacgc tcggtcgttc gactgcggcg agcggaaatg gcttacgaac ggggcggaga
     4621 tttcctggaa gatgccagga agatacttaa cagggaagtg agagggccgc ggcaaagccg
     4681 tttttccata ggctccgccc ccctgacaag catcacgaaa tctgacgctc aaatcagtgg
     4741 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggcgg ctccctcgtg
     4801 cgctctcctg ttcctgcctt tcggtttacc ggtgtcattc cgctgttatg gccgcgtttg
     4861 tctcattcca cgcctgacac tcagttccgg gtaggcagtt cgctccaagc tggactgtat
     4921 gcacgaaccc cccgttcagt ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     4981 caacccggaa agacatgcaa aagcaccact ggcagcagcc actggtaatt gatttagagg
     5041 agttagtctt gaagtcatgc gccggttaag gctaaactga aaggacaagt tttggtgact
     5101 gcgctcctcc aagccagtta cctcggttca aagagttggt agctcagaga accttcgaaa
     5161 aaccgccctg caaggcggtt ttttcgtttt cagagcaaga gattacgcgc agaccaaaac
     5221 gatctcaaga agatcatctt attaatcaga taaaatattt ctagatttca gtgcaattta
     5281 tctcttcaaa tgtagcacct gaagtcagcc ccataatcct tgttagatta tcaaagtttc
     5341 aaacgctaaa ttgcctgatg cgctacgctt atcaggccta catgatctct gcaatatatt
     5401 gagtttgcgt gcttttgtag gccggataag gcgttcacgc cgcatccggc aagaaacagc
     5461 aaacaatcca aaacgccgcg ttcagcggcg ttttttctgc ttttcttcgc gaattaattc
     5521 cgcttcgcaa catgtgagca ccggtttatt gactaccgga agcagtgtga ccgtgtgctt
     5581 ctcaaatgcc tgaggccagt ttgctcaggc tctccccgtg gaggtaataa ttgacgatat
     5641 gatcagtgca cggctaacta agcggcctgc tgactttctc gccgatcaaa aggcattttg
     5701 ctattaaggg attgacgagg gcgtatctgc gcagtaagat gcgccccgca ttccggcggt
     5761 agttcagcag ggcagaacgg cggactctaa atccgcatgg caggggttca aatcccctcc
     5821 gccggaccaa attcgaaaag cctgctcaac gagcaggctt ttttgcatgc tcgagcagct
     5881 cagggtcgaa tttgctttcg aatttctgcc attcatccgc ttattatcac ttattcaggc
     5941 gtagcaacca ggcgtttaag ggcaccaata actgccttaa aaaaattacg ccccgccctg
     6001 ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga agccatcaca
     6061 aacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt gcgtataata
     6121 tttgcccatg gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt ttaaatcaaa
     6181 actggtgaaa ctcacccagg gattggctga gacgaaaaac atattctcaa taaacccttt
     6241 agggaaatag gccaggtttt caccgtaaca cgccacatct tgcgaatata tgtgtagaaa
     6301 ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag tttgctcatg
     6361 gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt ctttcattgc
     6421 catacggaat tccggatgag cattcatcag gcgggcaaga atgtgaataa aggccggata
     6481 aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca gctgaacggt
     6541 ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt tacgatgcca
     6601 ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag cttccttagc
     6661 tcctgaaaat ctcgataact caaaaaatac gcccggtagt gatcttattt cattatggtg
     6721 aaagttggaa cctcttacgt gccgatcaac gtctcatttt cgccaaaagt tggcccaggg
     6781 cttcccggta tcaacaggga caccaggatt tatttattct gcgaagtgat cttccgtcac
     6841 aggtatttat tcggcgcaaa gtgcgtcggg tgatgctgcc aacttactga tttagtgtat
     6901 gatggtgttt ttgaggtgct ccagtggctt ctgtttctat cagctgtccc tcctgttcag
     6961 ctactgacgg ggtggtgcgt aacggcaaaa gcaccgccgg acatcagcgc tagcggagtg
     7021 tatactggct tactatgttg gcactgatga gggtgtcagt gaagtgcttc atgtggcagg
     7081 agaaaaaagg ctgcaccggt gcgtcagcag aatatgtgat acaggatata ttccgcttcc
     7141 tcgctcactg actcgctacg ctcggtcgtt cgactgcggc gagcggaaat ggcttacgaa
     7201 cggggcggag atttcctgga agatgccagg aagatactta acagggaagt gagagggccg
     7261 cggcaaagcc gtt
//
