LOCUS       sequence-351340-        5064 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tetrazine 3.0 tRNA synthetase and cognate amber suppressing tRNA 
            derived from M. barkeri for expression in E.coli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5064)
  AUTHORS   Jang HS, Jana S, Blizzard RJ, Meeuwsen JC, Mehl RA
  TITLE     Access to Faster Eukaryotic Cell Labeling with Encoded Tetrazine 
            Amino Acids.
  JOURNAL   J Am Chem Soc. 2020 Apr 22;142(16):7245-7249. doi: 
            10.1021/jacs.9b11520. Epub 2020 Apr 10.
  PUBMED    32251579
REFERENCE   2  (bases 1 to 5064)
  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/jacs.9b11520"; journalName: "J Am Chem Soc"; date: 
            "2020-04-22- 22"; volume: "142"; issue: "16"; pages: "7245-7249"
COMMENT     Sequence Label: pDule-Tet3.0
FEATURES             Location/Qualifiers
     source          1..5064
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1513..1542
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1558..1575
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1631..1648)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1801..1887
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(2094..2111)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2228..2773)
                     /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(2308..2327)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        join(2885..2890,2891..2907,2908..2913)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     CDS             2961..4151
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     complement(3313..3332)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     4458..4480
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4472..4489
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4473..4489
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gccgttatac gttgtttacg ctttgaggaa tcccatatga tggataaaaa accgctggat
       61 gtgctgatta gcgcgaccgg cctgtggatg agccgtaccg gcaccctgca taaaatcaaa
      121 catcatgaag tgagccgcag caaaatctat attgaaatgg cgtgcggcga tcatctggtg
      181 gtgaacaaca gccgtagctg ccgtaccgcg cgtgcgtttc gtcatcataa ataccgcaaa
      241 acctgcaaac gttgccgtgt gagcgatgaa gatatcaaca actttctgac ccgtagcacc
      301 gaaagcaaaa acagcgtgaa agtgcgtgtg gtgagcgcgc cgaaagtgaa aaaagcgatg
      361 ccgaaaagcg tgagccgtgc gccgaaaccg ctggaaaata gcgtgagcgc gaaagcgagc
      421 accaacacca gccgtagcgt tccgagcccg gcgaaaagca ccccgaacag cagcgttccg
      481 gcgtctgcgc cggcaccgag cctgacccgc agccagctgg atcgtgtgga agcgctgctg
      541 tctccggaag ataaaattag cctgaacatg gcgaaaccgt ttcgtgaact ggaaccggaa
      601 ctggtgaccc gtcgtaaaaa cgattttcag cgcctgtata ccaacgatcg tgaagattat
      661 ctgggcaaac tggaacgtga tatcaccaaa ttttttgtgg atcgcggctt tctggaaatt
      721 aaaagcccga ttctgattcc ggcggaatat gtggaacgta tgggcattaa caacgacacc
      781 gaactgagca aacaaatttt ccgcgtggat aaaaacctgt gcctgcgtcc gatgctggcc
      841 ccgaccggtt ataactatct gcgtaaactg gatcgtattc tgccgggtcc gatcaaaatt
      901 tttgaagtgg gcccgtgcta tcgcaaagaa agcgatggca aagaacacct ggaagaattc
      961 accatggttg gttttgctca aatgggcagc ggctgcaccc gtgaaaacct ggaagcgctg
     1021 atcaaagaat tcctggatta tctggaaatc gacttcgaaa ttgtgggcga tagctgcatg
     1081 gtgtatggcg ataccctgga tattatgcat ggcgatctgg aactgagcag cgcggtggtg
     1141 ggtccggtta gcctggatcg tgaatggggc attgataaac cgtggattgg cgcgggtttt
     1201 ggcctggaac gtctgctgaa agtgatgcat ggcttcaaaa acattaaacg tgcgagccgt
     1261 agcgaaagct actataacgg cattagcacg aacctgtaac tgcagtttca aacgctaaat
     1321 tgcctgatgc gctacgctta tcaggctaca tgatctctgc aatatattga gtttgcgtgc
     1381 ttttgtaggc cggataaggc gttcacgccg catccggcag gaaacagcaa acaatccaaa
     1441 acgccgcgtt cagcggcgtt ttttctgcgt ttcttcgcga attaattccg ctctcgagcg
     1501 aagcttgggc ccgaacaaaa actcatctca gaagaggatc tgaatagcgc cgtcgaccat
     1561 catcatcatc atcattgagt ttaaacggtc tccagcttgg ctgttttggc ggatgagaga
     1621 agattttcag cctgatacag attaaatcag aacgcagaag cggtctgata aaacagaatt
     1681 tgcctggcgg cagtagcgcg gtggtcccac ctgaccccat gccgaactca gaagtgaaac
     1741 gccgtagcgc cgatggtagt gtggggtctc cccatgcgag agtagggaac tgccaggcat
     1801 caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg
     1861 gtgaacgctc tcctgagtag gacaaatccg ccgggagctg tccctcctgt tcagctactg
     1921 acggggtggt gcgtaacggc aaaagcaccg ccggacatca gcgctagcgg agtgtatact
     1981 ggcttactat gttggcactg atgagggtgt cagtgaagtg cttcatgtgg caggagaaaa
     2041 aaggctgcac cggtgcgtca gcagaatatg tgatacagga tatattccgc ttcctcgctc
     2101 actgactcgc tacgctcggt cgttcgactg cggcgagcgg aaatggctta cgaacggggc
     2161 ggagatttcc tggaagatgc caggaagata cttaacaggg aagtgagagg gccgcggcaa
     2221 agccgttttt ccataggctc cgcccccctg acaagcatca cgaaatctga cgctcaaatc
     2281 agtggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggcggctccc
     2341 tcgtgcgctc tcctgttcct gcctttcggt ttaccggtgt cattccgctg ttatggccgc
     2401 gtttgtctca ttccacgcct gacactcagt tccgggtagg cagttcgctc caagctggac
     2461 tgtatgcacg aaccccccgt tcagtccgac cgctgcgcct tatccggtaa ctatcgtctt
     2521 gagtccaacc cggaaagaca tgcaaaagca ccactggcag cagccactgg taattgattt
     2581 agaggagtta gtcttgaagt catgcgccgg ttaaggctaa actgaaagga caagttttgg
     2641 tgactgcgct cctccaagcc agttacctcg gttcaaagag ttggtagctc agagaacctt
     2701 cgaaaaaccg ccctgcaagg cggttttttc gttttcagag caagagttta cgcgcagacc
     2761 aaaacgatct caagaagatc atcttattaa tcagataaaa tatttctaga tttcagtgca
     2821 atttatctct tcaaatgtag cacctgaagt cagccccata cgatataagt tgtaattctc
     2881 atgtttgaca gcttatcatc gataagcttt aatgcggtag tttatcacag ttaaattgct
     2941 aacgcagtca ggcaccgtgt atgaaatcta acaatgcgct catcgtcatc ctcggcaccg
     3001 tcaccctgga tgctgtaggc ataggcttgg ttatgccggt actgccgggc ctcttgcggg
     3061 atatcgtcca ttccgacagc atcgccagtc actatggcgt gctgctagcg ctatatgcgt
     3121 tgatgcaatt tctatgcgca cccgttctcg gagcactgtc cgaccgcttt ggccgccgcc
     3181 cagtcctgct cgcttcgcta cttggagcca ctatcgacta cgcgatcatg gcgaccacac
     3241 ccgtcctgtg gatcctctac gccggacgca tcgtggccgg catcaccggc gccacaggtg
     3301 cggttgctgg cgcctatatc gccgacatca ccgatgggga agatcgggct cgccacttcg
     3361 ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg ccccgtggcc gggggactgt
     3421 tgggcgccat ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc
     3481 tactactggg ctgcttccta atgcaggagt cgcataaggg agagcgtcga ccgatgccct
     3541 tgagagcctt caacccagtc agctccttcc ggtgggcgcg gggcatgact atcgtcgccg
     3601 cacttatgac tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca gcgctctggg
     3661 tcattttcgg cgaggaccgc tttcgctgga gcgcgacgat gatcggcctg tcgcttgcgg
     3721 tattcggaat cttgcacgcc ctcgctcaag ccttcgtcac tggtcccgcc accaaacgtt
     3781 tcggcgagaa gcaggccatt atcgccggca tggcggccga cgcgctgggc tacgtcttgc
     3841 tggcgttcgc gacgcgaggc tggatggcct tccccattat gattcttctc gcttccggcg
     3901 gcatcgggat gcccgcgttg caggccatgc tgtccaggca ggtagatgac gaccatcagg
     3961 gacagcttca aggatcgctc gcggctctta ccagcctaac ttcgatcatt ggaccgctga
     4021 tcgtcacggc gatttatgcc gcctcggcga gcacatggaa cgggttggca tggattgtag
     4081 gcgccgccct ataccttgtc tgcctccccg cgttgcgtcg cggtgcatgg agccgggcca
     4141 cctcgacctg aatggaagcc ggcggcacct cgctaacgga ttcaccactc caagaattgg
     4201 agccaatcaa ttcttgcgga gaactgtgaa tgcgcaaacc aacccttggc agaacatatc
     4261 catcgcgtcc gccatctcca gcagccgcac gcggcgcatc tcgggctcct tgcatgcacc
     4321 attccttgcg gcggcggtgc tcaacggcct caacctacta ctgggctgct tcctaatgca
     4381 ggagtcgcat aagggagagc gtctggcgaa agggggatgt gctgcaaggc gattaagttg
     4441 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg ccaagcttaa
     4501 aaaaaatcct tagctttcgc taagatctgc agtggcggaa accccgggaa tctaacccgg
     4561 ctgaacggat ttagagtcca ttcgatctac atgatcaggt tcccgcggcc gcgaattcag
     4621 cgttacaagt attacacaaa gttttttatg ttgagaatat ttttttgatg gggcgccact
     4681 tatttttgat cgttcgctca aagaagcggc gccagggttg tttttctttt caccagtgag
     4741 acgggcaaca gaacgccatg agcggcctca tttcttattc tgagttacaa cagtccgcac
     4801 cgctgccggt agctccttcc ggtgggcgcg gggcatgact atcgtcgccg cacttatgac
     4861 tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca gcgcccaaca gtcccccggc
     4921 cacggggcct gccaccatac ccacgccgaa acaagcgccc tgcaccatta tgttccggat
     4981 ctgcatcgca ggatgctgct ggctaccctg tggaacacct acggtacctc gggttgtcag
     5041 cctgtcccgc ttataagatc atac
//
