LOCUS Exported 4682 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expression plasmid encoding the evolved MjTyrRS variant Iodo.5RS for amber suppression with p-iodophenylalanine. ACCESSION . VERSION . KEYWORDS pDB070.iodo.5 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4682) 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 4682) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4682 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(147..164) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(281..826) /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(361..380) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(974..1597) /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 1505..1524 /label=Tet-R /note="Tetracycline resistance gene, reverse primer" protein_bind 1705..1723 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" promoter 1724..1778 /gene="tetR" /label=tetR/tetA promoters /note="overlapping promoters for bacterial tetR and tetA" protein_bind 1729..1747 /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O1 for the tetR and tetA genes" protein_bind 1759..1777 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" primer_bind complement(2749..2766) /label=pBAD Reverse /note="For vectors with E. coli araBAD promoter, reverse primer" terminator 2919..2965 /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" CDS complement(3675..4334) /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" primer_bind 4249..4268 /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" promoter complement(4335..4437) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" ORIGIN 1 ccgccggaca tcagcgctag cggagtgtat actggcttac tatgttggca ctgatgaggg 61 tgtcagtgaa gtgcttcatg tggcaggaga aaaaaggctg caccggtgcg tcagcagaat 121 atgtgataca ggatatattc cgcttcctcg ctcactgact cgctacgctc ggtcgttcga 181 ctgcggcgag cggaaatggc ttacgaacgg ggcggagatt tcctggaaga tgccaggaag 241 atacttaaca gggaagtgag agggccgcgg caaagccgtt tttccatagg ctccgccccc 301 ctgacaagca tcacgaaatc tgacgctcaa atcagtggtg gcgaaacccg acaggactat 361 aaagatacca ggcgtttccc cctggcggct ccctcgtgcg ctctcctgtt cctgcctttc 421 ggtttaccgg tgtcattccg ctgttatggc cgcgtttgtc tcattccacg cctgacactc 481 agttccgggt aggcagttcg ctccaagctg gactgtatgc acgaaccccc cgttcagtcc 541 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggaaag acatgcaaaa 601 gcaccactgg cagcagccac tggtaattga tttagaggag ttagtcttga agtcatgcgc 661 cggttaaggc taaactgaaa ggacaagttt tggtgactgc gctcctccaa gccagttacc 721 tcggttcaaa gagttggtag ctcagagaac cttcgaaaaa ccgccctgca aggcggtttt 781 ttcgttttca gagcaagaga ttacgcgcag accaaaacga tctcaagaag atcatcttat 841 taatcagata aaatatttct agatttcagt gcaatttatc tcttcaaatg tagcacctga 901 agtcagcccc atacgatata agttgtaatt ctcatgtttg acagcttatc atcgataaag 961 ctatctagaa agcttaagac ccactttcac atttaagttg tttttctaat ccgcatatga 1021 tcaattcaag gccgaataag aaggctggct ctgcaccttg gtgatcaaat aattcgatag 1081 cttgtcgtaa taatggcggc atactatcag tagtaggtgt ttccctttct tctttagcga 1141 cttgatgctc ttgatcttcc aatacgcaac ctaaagtaaa atgccccact gcgctgagtg 1201 catataatgc attctctagt gaaaaacctt gttggcataa aaaggctaat tgattttcga 1261 gagtttcata ctgtttttct gtaggccgtg tacctaaatg tacttttgct ccatcgcgat 1321 gacttagtaa agcacatcta aaacttttag cgttattacg taaaaaatct tgccagcttt 1381 ccccttctaa agggcaaaag tgagtatggt gcctatctaa catctcaatg gctaaggcgt 1441 cgagcaaagc ccgcttattt tttacatgcc aatacaatgt aggctgctct acacctagct 1501 tctgggcgag tttacgggtt gttaaacctt cgattccgac ctcattaagc agctctaatg 1561 cgctgttaat cactttactt ttatctaatc tagacatatg aattcggggc tttatgaatc 1621 tattatacag aaaaattttc ctgaaagcaa ataattaatt gcgttgcgca ttctagagca 1681 cagctccatc gaatggccag atgatcccta tcagtgatag agattgacac tctatcattg 1741 atagagttat tttaccactc cctatcagtg atagagaaag acctatggac gaatttgaaa 1801 tgataaagag aaacacatct gaaattatca gcgaggaaga gttaagagag gttttaaaaa 1861 aagatgaaaa atctgctctg ataggttttg aaccaagtgg taaaatacat ttagggcatt 1921 atctccaaat aaaaaagatg attgatttac aaaatgctgg atttgatata attatattgt 1981 tggctgattt ccacgcctat ttaaaccaga aaggagagtt ggatgagatt agaaaaatag 2041 gagattataa caaaaaagtt tttgaagcaa tggggttaaa ggcaaaatat gtttatggaa 2101 gttcgttcca gcttgataag gattatacac tgaatgtcta tagattggct ttaaaaacta 2161 ccttaaaaag agcaagaagg agtatggaac ttatagcaag agaggatgaa aatccaaagg 2221 ttgctgaagt tatctatcca ataatgcagg ttaatcctct taattatgag ggcgttgatg 2281 ttgcagttgg agggatggag cagagaaaaa tacacatgtt agcaagggag cttttaccaa 2341 aaaaggttgt ttgtattcac aaccctgtct taacgggttt ggatggagaa ggaaagatga 2401 gttcttcaaa agggaatttt atagctgttg atgactctcc agaagagatt agggctaaga 2461 taaagaaagc atactgccca gctggaattg ttgaaggaaa tccaataatg gagatagcta 2521 aatacttcct tgaatatcct ttaaccataa aaaggccaga aaaatttggt ggagatttga 2581 cagttaatag ctatgaggag ttagagagtt tatttaaaaa taaggaattg catccaatgc 2641 gcttaaaaaa tgctgtagct gaagaactta taaagatttt agagccaatt agaaagagat 2701 tataagtctc cagcttggct gttttggcgg atgagagaag attttcagcc tgatacagat 2761 taaatcagaa cgcagaagcg gtctgataaa acagaatttg cctggcggca gtagcgcggt 2821 ggtcccacct gaccccatgc cgaactcaga agtgaaacgc cgtagcgccg atggtagtgt 2881 ggggtctccc catgcgagag tagggaactg ccaggcatca aataaaacga aaggctcagt 2941 cgaaagactg ggcctttcgt tttatctgtt gtttgtgagc tcccggtcat caatcatccc 3001 cataatcctt gttagattat caaagtttca aacgctaaat tgcctgatgc gctacgctta 3061 tcaggcctac atgatctctg caatatattg agtttgcgtg cttttgtagg ccggataagg 3121 cgttcacgcc gcatccggca agaaacagca aacaatccaa aacgccgcgt tcagcggcgt 3181 tttttctgct tttcttcgcg aattaattcc gcttcgcaac atgtgagcac cggtttattg 3241 actaccggaa gcagtgtgac cgtgtgcttc tcaaatgcct gaggccagtt tgctcaggct 3301 ctccccgtgg aggtaataat tgacgatatg atcagtgcac ggctaactaa gcggcctgct 3361 gactttctcg ccgatcaaaa ggcattttgc tattaaggga ttgacgaggg cgtatctgcg 3421 cagtaagatg cgccccgcat tccggcggta gttcagcagg gcagaacggc ggactctaaa 3481 tccgcatggc aggggttcaa atcccctccg ccggaccaaa ttcgaaaagc ctgctcaacg 3541 agcaggcttt tttgcatgct cgagcagctc agggtcgaat ttgctttcga atttctgcca 3601 ttcatccgct tattatcact tattcaggcg tagcaaccag gcgtttaagg gcaccaataa 3661 ctgccttaaa aaaattacgc cccgccctgc cactcatcgc agtactgttg taattcatta 3721 agcattctgc cgacatggaa gccatcacaa acggcatgat gaacctgaat cgccagcggc 3781 atcagcacct tgtcgccttg cgtataatat ttgcccatgg tgaaaacggg ggcgaagaag 3841 ttgtccatat tggccacgtt taaatcaaaa ctggtgaaac tcacccaggg attggctgag 3901 acgaaaaaca tattctcaat aaacccttta gggaaatagg ccaggttttc accgtaacac 3961 gccacatctt gcgaatatat gtgtagaaac tgccggaaat cgtcgtggta ttcactccag 4021 agcgatgaaa acgtttcagt ttgctcatgg aaaacggtgt aacaagggtg aacactatcc 4081 catatcacca gctcaccgtc tttcattgcc atacggaatt ccggatgagc attcatcagg 4141 cgggcaagaa tgtgaataaa ggccggataa aacttgtgct tatttttctt tacggtcttt 4201 aaaaaggccg taatatccag ctgaacggtc tggttatagg tacattgagc aactgactga 4261 aatgcctcaa aatgttcttt acgatgccat tgggatatat caacggtggt atatccagtg 4321 atttttttct ccattttagc ttccttagct cctgaaaatc tcgataactc aaaaaatacg 4381 cccggtagtg atcttatttc attatggtga aagttggaac ctcttacgtg ccgatcaacg 4441 tctcattttc gccaaaagtt ggcccagggc ttcccggtat caacagggac accaggattt 4501 atttattctg cgaagtgatc ttccgtcaca ggtatttatt cggcgcaaag tgcgtcgggt 4561 gatgctgcca acttactgat ttagtgtatg atggtgtttt tgaggtgctc cagtggcttc 4621 tgtttctatc agctgtccct cctgttcagc tactgacggg gtggtgcgta acggcaaaag 4681 ca //