LOCUS       sequence-392917-        5311 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Machinery plasmid containing a single copy of the Mb Tet3.0RS and 
            cognate tRNA for incorporation of Tet3.0 at TAA codons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5311)
  AUTHORS   Bednar RM, Jana S, Kuppa S, Franklin R, Beckman J, Antony E, Cooley 
            RB, Mehl RA
  TITLE     Genetic Incorporation of Two Mutually Orthogonal Bioorthogonal Amino
            Acids That Enable Efficient Protein Dual-Labeling in Cells.
  JOURNAL   ACS Chem Biol. 2021 Nov 19;16(11):2612-2622. doi: 
            10.1021/acschembio.1c00649. Epub 2021 Sep 30.
  PUBMED    34590824
REFERENCE   2  (bases 1 to 5311)
  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: 
            "10.1021/acschembio.1c00649"; journalName: "ACS Chem Biol"; date: 
            "2021-11-19- 19"; volume: "16"; issue: "11"; pages: "2612-2622"
COMMENT     Sequence Label: pUltraI-Tet3.0[TAA]
FEATURES             Location/Qualifiers
     source          1..5311
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..1083
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(20..39)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     rep_origin      1293..2031
                     /direction=RIGHT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     promoter        2079..2170
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2171..2962
                     /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"
     regulatory      3145..3154
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     3773..3790
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     protein_bind    5134..5150
                     /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(5158..5164,5165..5180,5181..5186))
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     promoter        5234..5311
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt
       61 tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg
      121 gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc gggcaaacag
      181 tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc gcaaattgtc
      241 gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa
      301 cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt
      361 gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga agctgcctgc
      421 actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa cagtattatt
      481 ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt gggtcaccag
      541 caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg tctggctggc
      601 tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga aggcgactgg
      661 agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat cgttcccact
      721 gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc
      781 gggctgcgcg ttggtgcgga catctcggta gtgggatacg acgataccga agacagctca
      841 tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg gcaaaccagc
      901 gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca gctgttgccc
      961 gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc
     1021 gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag
     1081 tgagcgcaac gcaattaatg taagttagct cactcattag gcaccgggat ctcgaccgat
     1141 gcccttgaga gccttcaacc cagtcagctc cttccggtgg gcgcggggca tgactaacat
     1201 gagaattaca acttatatcg tatggggctg acttcaggtg ctacatttga agagataaat
     1261 tgcactgaaa tctagagcgg ttcagtagaa aagatcaaag gatcttcttg agatcctttt
     1321 tttctgcgcg taatcttttg ccctgtaaac gaaaaaacca cctggggagg tggtttgatc
     1381 gaaggttaag tcagttgggg aactgcttaa ccgtggtaac tggctttcgc agagcacagc
     1441 aaccaaatct gtccttccag tgtagccgga ctttggcgca cacttcaaga gcaaccgcgt
     1501 gtttagctaa acaaatcctc tgcgaactcc cagttaccaa tggctgctgc cagtggcgtt
     1561 ttaccgtgct tttccgggtt ggactcaagt gaacagttac cggataaggc gcagcagtcg
     1621 ggctgaacgg ggagttcttg cttacagccc agcttggagc gaacgaccta caccgagccg
     1681 agataccagt gtgtgagcta tgagaaagcg ccacacttcc cgtaagggag aaaggcggaa
     1741 caggtatccg gtaaacggca gggtcggaac aggagagcgc aagagggagc gacccgccgg
     1801 aaacggtggg gatctttaag tcctgtcggg tttcgcccgt actgtcagat tcatggttga
     1861 gcctcacggc tcccacagat gcaccggaaa agcgtctgtt tatgtgaact ctggcaggag
     1921 ggcggagcct atggaaaaac gccaccggcg cggccctgct gttttgcctc acatgttagt
     1981 cccctgctta tccacggaat ctgtgggtaa ctttgtatgt gtccgcagcg cccgccgcag
     2041 tctcacgccc ggagcgtagc gaccgagtga gctagctatt tgtttatttt tctaaataca
     2101 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     2161 aaggaagagt atgagggaag cggtgatcgc cgaagtatcg actcaactat cagaggtagt
     2221 tggcgtcatc gagcgccatc tcgaaccgac gttgctggcc gtacatttgt acggctccgc
     2281 agtggatggc ggcctgaagc cacacagtga tattgatttg ctggttacgg tgaccgtaag
     2341 gcttgatgaa acaacgcggc gagctttgat caacgacctt ttggaaactt cggcttcccc
     2401 tggagagagc gagattctcc gcgctgtaga agtcaccatt gttgtgcacg acgacatcat
     2461 tccgtggcgt tatccagcta agcgcgaact gcaatttgga gaatggcagc gcaatgacat
     2521 tcttgcaggt atcttcgagc cagccacgat cgacattgat ctggctatct tgctgacaaa
     2581 agcaagagaa catagcgttg ccttggtagg tccagcggcg gaggaactct ttgatccggt
     2641 tcctgaacag gatctatttg aggcgctaaa tgaaacctta acgctatgga actcgccgcc
     2701 cgactgggct ggcgatgagc gaaatgtagt gcttacgttg tcccgcattt ggtacagcgc
     2761 agtaaccggc aaaatcgcgc cgaaggatgt cgctgccgac tgggcaatgg agcgcctgcc
     2821 ggcccagtat cagcccgtca tacttgaagc tagacaggct tatcttggac aagaagaaga
     2881 tcgcttggcc tcgcgcgcag atcagttgga agaatttgtc cactacgtga aaggcgagat
     2941 caccaaggta gtcggcaaat aatgtctaac aattcgttca agccgagggg ccgcaagatc
     3001 cggccacgat gacccggtcg tcggttcagg gcagggtcgt taaatagccg cttatgtcta
     3061 ttgctggttt accggtttat tgactaccgg aagcagtgtg accgtgtgct tctcaaatgc
     3121 ctgaggcgcc ggtggtacct ggtggccgcc atggcaaatt cgaccctgag ctgctcgagc
     3181 atgcaaaaaa gcctgctcgt tgagcaggct tttcgaattt ggcggaaacc ccgggaatct
     3241 aacccggctg aacggattta aagtccgttc gatctacatg atcaggtttc caatgcgggg
     3301 cgcatcttac tgcgcagata cgccctcgtc aatcccttaa tagcaaaatg ccttttgatc
     3361 ggcgagaaag tcagcaggcc gcttagttag ccgtgcactg atcatatcgt caattattac
     3421 ctccacgggg agagcctgag caaactggcc tcaggcattt gagaagcaca cggtcacact
     3481 gcttccggta gtcaataaac cggtgctcac atgttgcgaa gcggaattac ctgcaggcta
     3541 acaaggatta tggggatgat tgatgaccgg gagctcacaa acaaggccca gtctttcgac
     3601 tgagcctttc gttttatttg atgcctggca gttccctact ctcgcatggg gagaccccac
     3661 actaccatcg gcgctacggc gtttcacttc tgagttcggc atggggtcag gtgggaccac
     3721 cgcgctactg ccgccaggca aattctgttt tatcagaccg cttctgcgtt ctgatttaat
     3781 ctgtatcagg ctgaaaatct tctctcatcc gccaaaacag ccaagctgga gaccgtttaa
     3841 acgcggccgc tcacaggttc gtgctaatgc cgttatagta gctttcgcta cggctcgcac
     3901 gtttaatgtt tttgaagcca tgcatcactt tcagcagacg ttccaggcca aaacccgcgc
     3961 caatccacgg tttatcaatg ccccattcac gatccaggct aaccggaccc accaccgcgc
     4021 tgctcagttc cagatcgcca tgcataatat ccagggtatc gccatacacc atgcagctat
     4081 cgcccacaat ttcgaagtcg atttccagat aatccaggaa ttctttgatc agcgcttcca
     4141 ggttttcacg ggtgcagccg ctgcccattt gagcaaaacc aaccatggtg aattcttcca
     4201 ggtgttcttt gccatcgctt tctttgcgat agcacgggcc cacttcaaaa attttgatcg
     4261 gacccggcag aatacgatcc agtttacgca gatagttata accggtcggg gccagcatcg
     4321 gacgcaggca caggttttta tccacgcgga aaatttgttt gctcagttcg gtgtcgttgt
     4381 taatgcccat acgttccaca tattccgccg gaatcagaat cgggctttta atttccagaa
     4441 agccgcgatc cacaaaaaat ttggtgatat cacgttccag tttgcccaga taatcttcac
     4501 gatcgttggt atacaggcgc tgaaaatcgt ttttacgacg ggtcaccagt tccggttcca
     4561 gttcacgaaa cggtttcgcc atgttcaggc taattttatc ttccggagac agcagcgctt
     4621 ccacacgatc cagctggctg cgggtcaggc tcggtgccgg cgcagacgcc ggaacgctgc
     4681 tgttcggggt gcttttcgcc gggctcggaa cgctacggct ggtgttggtg ctcgctttcg
     4741 cgctcacgct attttccagc ggtttcggcg cacggctcac gcttttcggc atcgcttttt
     4801 tcactttcgg cgcgctcacc acacgcactt tcacgctgtt tttgctttcg gtgctacggg
     4861 tcagaaagtt gttgatatct tcatcgctca cacggcaacg tttgcaggtt ttgcggtatt
     4921 tatgatgacg aaacgcacgc gcggtacggc agctacggct gttgttcacc accagatgat
     4981 cgccgcacgc catttcaata tagattttgc tgcggctcac ttcatgatgt ttgattttat
     5041 gcagggtgcc ggtacggctc atccacaggc cggtcgcgct aatcagcaca tccagcggtt
     5101 ttttatccat gcggccgcac ctcctttgtg aaattgttat ccgctcacaa ttccacacat
     5161 tatacgagcc gatgattaat tgtcaacagc tccctaatgc aggagtcgca taagggagag
     5221 cgtcgagatc ccggacacca tcgaatggcg caaaaccttt cgcggtatgg catgatagcg
     5281 cccggaagag agtcaattca gggtggtgaa t
//
