LOCUS       sequence-346623-        3274 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  tRNA synthetase/tRNA pair for the in vivo incorporation of 
            L-3-(2-cyano-4-pyridyl)alanine (mCNP), 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 3274)
  AUTHORS   Abdelkader EH, Qianzhu H, George J, Frkic RL, Jackson CJ, Nitsche C,
            Otting G, Huber T
  TITLE     Genetic Encoding of Cyanopyridylalanine for In-Cell Protein 
            Macrocyclization by the Nitrile-Aminothiol Click Reaction.
  JOURNAL   Angew Chem Int Ed Engl. 2022 Jan 31. doi: 10.1002/anie.202114154.
  PUBMED    35102680
REFERENCE   2  (bases 1 to 3274)
  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. 2022 Jan 31. doi: 10.1002/anie.202114154."
COMMENT     Sequence Label: pRSF-G1mCNPRS
FEATURES             Location/Qualifiers
     source          1..3274
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(84..175)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     258..277
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        258..276
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             308..330
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(1235..1253)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1249..1296
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      complement(1777..2526)
                     /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(1787..1804)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2027..2046)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2542..3274,1..83))
                     /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(join(2542..3274,1..83))
                     /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     join(3265..3274,1..10)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 catttatacc catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac
       61 gtttcccgtt gaatatggct catactcttc ctttttcaat attattgaag catttatcag
      121 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggc
      181 atgcagcgct tgacttgacg ggacggcgtt gtaattctca tgtttgacag cttatcatcg
      241 ataagcttgg tacccaataa tacgactcac tatagggaga ccacaacggt ttccctctag
      301 aaataatttt gtttaacttt aagaaggaga tatacatatg gtggtgaaat ttaccgatag
      361 ccagattcag catctgatgg aatatggtga taatgattgg agcgaagccg aatttgaaga
      421 tgcagcagca cgtgataaag aatttagcag ccagtttagc aaactgaaaa gcgccaatga
      481 taaaggcctg aaagatgtta ttgcaaatcc gcgtaatgat ctgaccgatc tggaaaacaa
      541 aattcgcgaa aaactggcag cccgtggttt tattgaagtt cataccccga tttttgtgag
      601 caaaagcgca ctggcaaaaa tgaccattac cgaagatcat ccgctgttca aacaggtgtt
      661 ttggattgat gataaacgtg cactgcgtcc gatgcatgca atgaatgctt tgaaagttat
      721 gcgtgaactg cgcgatcata ccaaaggtcc ggttaaaatc tttgaaattg gtagctgctt
      781 tcgcaaagaa agcaaaagca gtacccatct ggaagaattt accatgctga acctggcaga
      841 aatgggtcct gatggtgatc cgatggaaca tctgaaaatg tatattggcg atatcatgga
      901 tgccgttggt gttgaatata ccaccagtcg tgaagaatca gatgtttggg ttgaaaccct
      961 ggacgtggaa attaatggca ccgaagttgc aagcggttct gttggtccgc ataaactgga
     1021 tccggcacat gatgtgcatg aaccgtgggc aggtattggt tttggtctgg aacgtctgct
     1081 gatgctgaaa aatggtaaaa gcaatgcacg caaaaccggc aaaagtatta cctatctgaa
     1141 tggctacaaa ctggattaag aattcgagct cccgggtacc agatagatcc ggctgctaac
     1201 aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc
     1261 cttggggcct ctaaacgggt cttgaggggt tttttggttt gtgagctccc gtaattccgc
     1321 ttcgcaacat gtgagcaccg gtttattgac taccggaagc agtgtgaccg tgtgcttctc
     1381 aaatgcctga ggccagtttg ctcaggctct ccccgtggag gtaataattg acgatatgat
     1441 cagtgcacgg ctaactaagc ggcctgctga ctttctcgcc gatcaaaagg cattttgcta
     1501 ttaagggatt gacgagggcg tatctgcgca gtaagatgcg ccccgcattg gagggcgctc
     1561 cggcgagcaa acgggtctct aaaacctgta agcggggttc gaccccccgg cctttcgcca
     1621 aattcgaaaa gcctgctcaa cgagcaggct tttttgcatg ctcgagcagc tcagggtcga
     1681 atttgctttc gaatttctgc cattcatccg cttattatca cttattcagg cgtagcaacc
     1741 aggcgtttaa gggcaccaat aactgcctta aaaaaacttc cgcttcctcg ctcactgact
     1801 cgctacgctc ggtcgttcga ctgcggcgag cggtgtcagc tcactcaaaa gcggtaatac
     1861 ggttatccac agaatcaggg gataaagccg gaaagaacat gtgagcaaaa agcaaagcac
     1921 cggaagaagc caacgccgca ggcgtttttc cataggctcc gcccccctga cgagcatcac
     1981 aaaaatcgac gctcaagcca gaggtggcga aacccgacag gactataaag ataccaggcg
     2041 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     2101 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgttggtat
     2161 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     2221 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     2281 ttatcgccac tggcagcagc cattggtaac tgatttagag gactttgtct tgaagttatg
     2341 cacctgttaa ggctaaactg aaagaacaga ttttggtgag tgcggtcctc caacccactt
     2401 accttggttc aaagagttgg tagctcagcg aaccttgaga aaaccaccgt tggtagcggt
     2461 ggtttttctt tatttatgag atgatgaatc aatcggtcta tcaagtcaac gaacagctat
     2521 tccgttgaac tgtggtacgc gttagaaaaa ctcatcgagc atcaaatgaa actgcaattt
     2581 attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga
     2641 aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac
     2701 tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga
     2761 gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagtttat gcatttcttt
     2821 ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa
     2881 accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcggtcgc tgttaaaagg
     2941 acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat
     3001 attttcacct gaatcaggat attcttctaa tacctggaat gctgttttcc cggggatcgc
     3061 agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg
     3121 cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct
     3181 acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca atcgatagat
     3241 tgtcgcacct gattgcccga cattatcgcg agcc
//
