LOCUS       sequence-459819-        3390 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  tRNA synthetase/tRNA pair for the in vivo incorporation of 
            (7-hydroxy-4-coumarin-yl) ethylglycine (Hco), into proteins in E. 
            coli in response to the amber (TAG) codon.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3390)
  AUTHORS   Qianzhu H, Abdelkader EH, Welegedara AP, Habel E, Paul N, Frkic RL, 
            Jackson CJ, Huber T, Otting G
  TITLE     Rendering Proteins Fluorescent Inconspicuously: Genetically Encoded 
            4-Cyanotryptophan Conserves Their Structure and Enables the 
            Detection of Ligand Binding Sites.
  JOURNAL   Angew Chem Int Ed Engl. 2024 Dec 4:e202421000. doi: 
            10.1002/anie.202421000.
  PUBMED    39632265
REFERENCE   2  (bases 1 to 3390)
  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: "Angew Chem 
            Int Ed Engl. 2024 Dec 4:e202421000. doi: 10.1002/anie.202421000."
COMMENT     Sequence Label: pRSF-MjCouRS
FEATURES             Location/Qualifiers
     source          1..3390
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     6..25
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6..24
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             56..78
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(1093..1111)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1107..1154
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      complement(1641..2390)
                     /direction=LEFT
                     /label=RSF ori
                     /note="Plasmids containing the RSF 1030 origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     primer_bind     complement(1651..1668)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(1891..1910)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2406..3221)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2406..3221)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3129..3148
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3222..3313)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cccaataata cgactcacta tagggagacc acaacggttt ccctctagaa ataattttgt
       61 ttaactttaa gaaggagata tacatatgga cgaatttgaa atgataaaga gaaacacatc
      121 tgaaattatc agcgaggaag agttaagaga ggttttaaaa aaagatgaaa aatctgctga
      181 gataggtttt gaaccaagtg gtaaaataca tttagggcat tatctccaaa taaaaaagat
      241 gattgattta caaaatgctg gatttgatat aattatacat ttgggtgatt taggcgccta
      301 tttaaaccag aaaggagagt tggatgagat tagaaaaata ggagattata acaaaaaagt
      361 ttttgaagca atggggttaa aggcaaaata tgtttatgga agtgaatatc atcttgataa
      421 ggattataca ctgaatgtct atagattggc tttaaaaact accttaaaaa gagcaagaag
      481 gagtatggaa cttatagcaa gagaggatga aaatccaaag gttgctgaag ttatctatcc
      541 aataatgcag gttaatggga ttcattatgg tggcgttgat gttgcagttg gagggatgga
      601 gcagagaaaa atacacatgt tagcaaggga gcttttacca aaaaaggttg tttgtattca
      661 caaccctgtc ttaacgggtt tggatggaga aggaaagatg agttcttcaa aagggaattt
      721 tatagctgtt gatgactctc cagaagagat tagggctaag ataaagaaag catactgccc
      781 agctggagtt gttgaaggaa atccaataat ggagatagct aaatacttcc ttgaatatcc
      841 tttaaccata aaaaggccag aaaaatttgg tggagatttg acagttaata gctatgagga
      901 gttagagagt ttatttaaaa ataaggaatt gcatccaatg gatttaaaaa atgctgtagc
      961 tgaagaactt ataaagattt tagagccaat tagaaagaga ttataagaat tcgagctccc
     1021 gggtaccatg gcatgcatcg atagatccgg ctgctaacaa agcccgaaag gaagctgagt
     1081 tggctgctgc caccgctgag caataactag cataacccct tggggcctct aaacgggtct
     1141 tgaggggttt tttggtttgt gagctcccgt aattccgctt cgcaacatgt gagcaccggt
     1201 ttattgacta ccggaagcag tgtgaccgtg tgcttctcaa atgcctgagg ccagtttgct
     1261 caggctctcc ccgtggaggt aataattgac gatatgatca gtgcacggct aactaagcgg
     1321 cctgctgact ttctcgccga tcaaaaggca ttttgctatt aagggattga cgagggcgta
     1381 tctgcgcagt aagatgcgcc ccgcattccg gcggtagttc agcagggcag aacggcggac
     1441 tctaaatccg catggcaggg gttcaaatcc cctccgccgg accaaattcg aaaagcctgc
     1501 tcaacgagca ggcttttttg catgctcgag cagctcaggg tcgaatttgc tttcgaattt
     1561 ctgccattca tccgcttatt atcacttatt caggcgtagc aaccaggcgt ttaagggcac
     1621 caataactgc cttaaaaaaa cttccgcttc ctcgctcact gactcgctac gctcggtcgt
     1681 tcgactgcgg cgagcggtgt cagctcactc aaaagcggta atacggttat ccacagaatc
     1741 aggggataaa gccggaaaga acatgtgagc aaaaagcaaa gcaccggaag aagccaacgc
     1801 cgcaggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     1861 gccagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     1921 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     1981 cttcgggaag cgtggcgctt tctcatagct cacgctgttg gtatctcagt tcggtgtagg
     2041 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     2101 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     2161 cagccattgg taactgattt agaggacttt gtcttgaagt tatgcacctg ttaaggctaa
     2221 actgaaagaa cagattttgg tgagtgcggt cctccaaccc acttaccttg gttcaaagag
     2281 ttggtagctc agcgaacctt gagaaaacca ccgttggtag cggtggtttt tctttattta
     2341 tgagatgatg aatcaatcgg tctatcaagt caacgaacag ctattccgtt gaactgtggt
     2401 acgcgttaga aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta
     2461 tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag
     2521 ttccatagga tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata
     2581 caacctatta atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg
     2641 acgactgaat ccggtgagaa tggcaaaagt ttatgcattt ctttccagac ttgttcaaca
     2701 ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt
     2761 gattgcgcct gagcgagacg aaatacgcgg tcgctgttaa aaggacaatt acaaacagga
     2821 atcgaatgca accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca
     2881 ggatattctt ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat
     2941 gcatcatcag gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc
     3001 cagtttagtc tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc
     3061 agaaacaact ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc
     3121 ccgacattat cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat
     3181 cgcggcctag agcaagacgt ttcccgttga atatggctca tactcttcct ttttcaatat
     3241 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     3301 aaaaataaac aaataggcat gcagcgcttg acttgacggg acggcgttgt aattctcatg
     3361 tttgacagct tatcatcgat aagcttggta
//
