LOCUS       sequence-491665-        7858 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  L-arabinose inducible E.coli cascade operon inserted in pCDF vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7858)
  AUTHORS   Lee C, Kim S
  TITLE     Continuous targeted hypermutation with a tunable mutation window.
  JOURNAL   Nat Commun. 2025 Nov 25. doi: 10.1038/s41467-025-66736-2.
  PUBMED    41290661
REFERENCE   2  (bases 1 to 7858)
  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: "Nat Commun.
            2025 Nov 25. doi: 10.1038/s41467-025-66736-2."
COMMENT     Sequence Label: pCDF-cascade
FEATURES             Location/Qualifiers
     source          1..7858
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(129..1007)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        1034..1318
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     1241..1260
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             5718..5762
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(5800..5818)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5814..5861
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(6032..6823)
                     /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"
     promoter        complement(6824..6915)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6963..7701)
                     /direction=LEFT
                     /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."
ORIGIN
        1 tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc gcgtcaaatg
       61 gacgaagcag ggattctgca aaccctatgc tactccgtca agccgtcaat tgtctgattc
      121 gttaccaatt atgacaactt gacggctaca tcattcactt tttcttcaca accggcacgg
      181 aactcgctcg ggctggcccc ggtgcatttt ttaaataccc gcgagaaata gagttgatcg
      241 tcaaaaccaa cattgcgacc gacggtggcg ataggcatcc gggtggtgct caaaagcagc
      301 ttcgcctggc tgatacgttg gtcctcgcgc cagcttaaga cgctaatccc taactgctgg
      361 cggaaaagat gtgacagacg cgacggcgac aagcaaacat gctgtgcgac gctggcgata
      421 tcaaaattgc tgtctgccag gtgatcgctg atgtactgac aagcctcgcg tacccgatta
      481 tccatcggtg gatggagcga ctcgttaatc gcttccatgc gccgcagtaa caattgctca
      541 agcagattta tcgccagcag ctccgaatag cgcccttccc cttgcccggc gttaatgatt
      601 tgcccaaaca ggtcgctgaa atgcggctgg tgcgcttcat ccgggcgaaa gaaccccgta
      661 ttggcaaata ttgacggcca gttaagccat tcatgccagt aggcgcgcgg acgaaagtaa
      721 acccactggt gataccattc gcgagcctcc ggatgacgac cgtagtgatg aatctctcct
      781 ggcgggaaca gcaaaatatc acccggtcgg caaacaaatt ctcgtccctg atttttcacc
      841 accccctgac cgcgaatggt gagattgaga atataacctt tcattcccag cggtcggtcg
      901 ataaaaaaat cgagataacc gttggcctca atcggcgtta aacccgccac cagatgggca
      961 ttaaacgagt atcccggcag caggggatca ttttgcgctt cagccatact tttcatactc
     1021 ccgccattca gagaagaaac caattgtcca tattgcatca gacattgccg tcactgcgtc
     1081 ttttactggc tcttctcgct aaccaaaccg gtaaccccgc ttattaaaag cattctgtaa
     1141 caaagcggga ccaaagccat gacaaaaacg cgtaacaaaa gtgtctataa tcacggcaga
     1201 aaagtccaca ttgattattt gcacggcgtc acactttgct atgccatagc atttttatcc
     1261 ataagattag cggatcctac ctgacgcttt ttatcgcaac tctctactgt ttctccatac
     1321 gaaatgcagg agaaatgtcg gagcatatga aggagaacaa atgaatttgc ttattgataa
     1381 ctggatccct gtacgcccgc gaaacggggg gaaagtccaa atcataaatc tgcaatcgct
     1441 atactgcagt agagatcagt ggcgattaag tttgccccgt gacgatatgg aactggccgc
     1501 tttagcactg ctggtttgca ttgggcaaat tatcgccccg gcaaaagatg acgttgaatt
     1561 tcgacatcgc ataatgaatc cgctcactga agatgagttt caacaactca tcgcgccgtg
     1621 gatagatatg ttctacctta atcacgcaga acatcccttt atgcagacca aaggtgtcaa
     1681 agcaaatgat gtgactccaa tggaaaaact gttggctggg gtaagcggcg cgacgaattg
     1741 tgcatttgtc aatcaaccgg ggcagggtga agcattatgt ggtggatgca ctgcgattgc
     1801 gttattcaac caggcgaatc aggcaccagg ttttggtggt ggttttaaaa gcggtttacg
     1861 tggaggaaca cctgtaacaa cgttcgtacg tgggatcgat cttcgttcaa cggtgttact
     1921 caatgtcctc acattacctc gtcttcaaaa acaatttcct aatgaatcac atacggaaaa
     1981 ccaacctacc tggattaaac ctatcaagtc caatgagtct atacctgctt cgtcaattgg
     2041 gtttgtccgt ggtctattct ggcaaccagc gcatattgaa ttatgcgatc ccattgggat
     2101 tggtaaatgt tcttgctgtg gacaggaaag caatttgcgt tataccggtt ttcttaagga
     2161 aaaatttacc tttacagtta atgggctatg gccccatccg cattcccctt gtctggtaac
     2221 agtcaagaaa ggggaggttg aggaaaaatt tcttgctttc accacctccg caccatcatg
     2281 gacacaaatc agccgagttg tggtagataa gattattcaa aatgaaaatg gaaatcgcgt
     2341 ggcggcggtt gtgaatcaat tcagaaatat tgcgccgcaa agtcctcttg aattgattat
     2401 ggggggatat cgtaataatc aagcatctat tcttgaacgg cgtcatgatg tgttgatgtt
     2461 taatcagggg tggcaacaat acggcaatgt gataaacgaa atagtgactg ttggtttggg
     2521 atataaaaca gccttacgca aggcgttata tacctttgca gaagggttta aaaataaaga
     2581 cttcaaaggg gccggagtct ctgttcatga gactgcagaa aggcatttct atcgacagag
     2641 tgaattatta attcccgatg tactggcgaa tgttaatttt tcccaggctg atgaggtaat
     2701 agctgattta cgagacaaac ttcatcaatt gtgtgaaatg ctatttaatc aatctgtagc
     2761 tccctatgca catcatccta aattaataag cacattagcg cttgcccgcg ccacgctata
     2821 caaacattta cgggagttaa aaccgcaagg agggccatca aatggctgat gaaattgatg
     2881 caatggcttt atatcgagcc tggcaacaac tggataatgg atcatgtgcg caaattagac
     2941 gtgtttcaga acctgatgaa ttacgcgata tccctgcgtt ttataggctg gtgcaacctt
     3001 ttggttggga aaacccacgt caccagcagg ctcttttgcg catggtgttt tgcctgagcg
     3061 caggaaagaa tgtcatccga catcaggaca aaaaatcgga gcaaacaaca ggtatctcgt
     3121 tgggaagagc tttagccaat agtggaagaa ttaacgagcg ccgtatcttt caattaattc
     3181 gggctgacag aacagccgat atggtccagt tacgtcgatt acttactcac gccgaacccg
     3241 tacttgactg gccattaatg gccaggatgt tgacctggtg gggaaagcgc gaacgccagc
     3301 aacttctgga agattttgta ttgaccacaa acaaaaatgc gtaaggaaac ctttctatgt
     3361 ctaactttat caatattcat gttctgatct ctcacagccc ttcatgtctg aaccgcgacg
     3421 atatgaacat gcagaaagac gctattttcg gcggcaaaag acgagtaaga atttcaagtc
     3481 aaagccttaa acgtgcgatg cgtaaaagtg gttattacgc acaaaatatt ggtgaatcca
     3541 gtctcagaac cattcatctt gcacaattac gtgatgttct tcggcaaaaa cttggtgaac
     3601 gttttgacca aaaaatcatc gataagacat tagcgctgct ctccggtaaa tcagttgatg
     3661 aagccgaaaa gatttctgcc gatgcggtta ctccctgggt tgtgggagaa atagcctggt
     3721 tctgtgagca ggttgcaaaa gcagaggctg ataatctgga tgataaaaag ctgctcaaag
     3781 ttcttaagga agatattgcc gccatacgtg tgaatttaca gcagggtgtt gatattgcgc
     3841 ttagtggaag aatggcaacc agcggcatga tgactgagtt gggaaaagtt gatggtgcaa
     3901 tgtccattgc gcatgcgatc actactcatc aggttgattc tgatattgac tggttcaccg
     3961 ctgtagatga tttacaggaa caaggttctg cacatctggg aactcaggaa ttttcatcgg
     4021 gtgtttttta tcgttatgcc aacattaacc tcgctcaact tcaggaaaat ttaggtggtg
     4081 cctccaggga gcaggctctg gaaattgcaa cccatgttgt tcatatgctg gcaacagagg
     4141 tccctggagc aaaacagcgt acttatgccg cttttaaccc tgcggatatg gtaatggtta
     4201 atttctccga tatgccactt tctatggcaa atgcttttga aaaagcggtt aaagcgaaag
     4261 atggcttttt gcaaccgtct atacaggcgt ttaatcaata ttgggatcgc gttgccaatg
     4321 gatatggtct gaacggagct gctgcgcaat tcagcttatc tgatgtagac ccaattactg
     4381 ctcaagttaa acaaatgcct actttagaac agttaaaatc ctgggttcgt aataatggcg
     4441 aggcgtgaac atgagatctt atttgatctt gcggcttgct gggccaatgc aagcctgggg
     4501 gcagccgacc tttgaaggaa cgcgacctac cggaagattt ccgacccgaa gcgggttatt
     4561 agggctactc ggggcttgtc ttgggatcca acgtgatgat acttcttcat tacaggcgtt
     4621 atcagagagt gtgcaatttg cagtgcgctg cgatgaactc attcttgacg atcgtcgtgt
     4681 gtctgtaacg gggttgcgtg attaccatac agtccttgga gcgcgagaag attaccgtgg
     4741 tttgaaaagt catgaaacga ttcaaacatg gcgcgaatat ttatgtgatg cctcctttac
     4801 cgtcgctctc tggttaacac cccatgcaac gatggttatc tcagaacttg aaaaagcagt
     4861 attaaagcct cggtatacac cttacctggg gcggagaagt tgcccactaa cacacccgct
     4921 ttttttgggg acatgtcagg catcggatcc tcagaaggcg ctattaaatt atgagcccgt
     4981 tggcggcgat atatatagtg aggaatcagt tacagggcat catttaaaat ttacggcgcg
     5041 cgacgaaccg atgatcacct tgcctcgaca atttgcttcc cgagaatggt atgtgattaa
     5101 aggaggtatg gatgtatctc agtaaagtca tcattgccag ggcctggagc agggatcttt
     5161 accaacttca ccagggatta tggcatttat ttccaaacag accggatgct gctcgtgatt
     5221 ttctttttca tgttgagaag cgaaacacac cagaaggctg tcatgtttta ttgcagtcag
     5281 cgcaaatgcc tgtttcaact gccgttgcga cagtcattaa aactaaacag gttgaatttc
     5341 aacttcaggt tggtgttcca ctctattttc ggcttcgggc aaatccgatc aaaactattc
     5401 tcgacaatca aaagcgcctg gacagtaaag ggaatattaa acgctgtcgg gttccgttaa
     5461 taaaagaagc agaacaaatc gcgtggttgc aacgtaaatt gggcaatgcg gcgcgcgttg
     5521 aagatgtgca tcccatatcg gaacggccac agtatttttc tggtgatggt aaaagtggaa
     5581 agatccaaac ggtttgcttt gaaggtgtgc tcaccatcaa cgacgcgcca gcgttaatag
     5641 atcttgtaca gcaaggtatt gggccagcta aatcgatggg atgtggcttg ctatctttgg
     5701 ctccactgtg ataacgtaaa gaaaccgctg ctgcgaaatt tgaacgccag cacatggact
     5761 cgtctactag cgcagcttaa ttaacctagg ctgctgccac cgctgagcaa taactagcat
     5821 aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaacct caggcatttg
     5881 agaagcacac ggtcacactg cttccggtag tcaataaacc ggtaaaccag caatagacat
     5941 aagcggctat ttaacgaccc tgccctgaac cgacgaccgg gtcatcgtgg ccggatcttg
     6001 cggcccctcg gcttgaacga attgttagac attatttgcc gactaccttg gtgatctcgc
     6061 ctttcacgta gtggacaaat tcttccaact gatctgcgcg cgaggccaag cgatcttctt
     6121 cttgtccaag ataagcctgt ctagcttcaa gtatgacggg ctgatactgg gccggcaggc
     6181 gctccattgc ccagtcggca gcgacatcct tcggcgcgat tttgccggtt actgcgctgt
     6241 accaaatgcg ggacaacgta agcactacat ttcgctcatc gccagcccag tcgggcggcg
     6301 agttccatag cgttaaggtt tcatttagcg cctcaaatag atcctgttca ggaaccggat
     6361 caaagagttc ctccgccgct ggacctacca aggcaacgct atgttctctt gcttttgtca
     6421 gcaagatagc cagatcaatg tcgatcgtgg ctggctcgaa gatacctgca agaatgtcat
     6481 tgcgctgcca ttctccaaat tgcagttcgc gcttagctgg ataacgccac ggaatgatgt
     6541 cgtcgtgcac aacaatggtg acttctacag cgcggagaat ctcgctctct ccaggggaag
     6601 ccgaagtttc caaaaggtcg ttgatcaaag ctcgccgcgt tgtttcatca agccttacgg
     6661 tcaccgtaac cagcaaatca atatcactgt gtggcttcag gccgccatcc actgcggagc
     6721 cgtacaaatg tacggccagc aacgtcggtt cgagatggcg ctcgatgacg ccaactacct
     6781 ctgatagttg agtcgatact tcggcgatca ccgcttccct catactcttc ctttttcaat
     6841 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6901 agaaaaataa acaaatagct agctcactcg gtcgctacgc tccgggcgtg agactgcggc
     6961 gggcgctgcg gacacataca aagttaccca cagattccgt ggataagcag gggactaaca
     7021 tgtgaggcaa aacagcaggg ccgcgccggt ggcgtttttc cataggctcc gccctcctgc
     7081 cagagttcac ataaacagac gcttttccgg tgcatctgtg ggagccgtga ggctcaacca
     7141 tgaatctgac agtacgggcg aaacccgaca ggacttaaag atccccaccg tttccggcgg
     7201 gtcgctccct cttgcgctct cctgttccga ccctgccgtt taccggatac ctgttccgcc
     7261 tttctccctt acgggaagtg tggcgctttc tcatagctca cacactggta tctcggctcg
     7321 gtgtaggtcg ttcgctccaa gctgggctgt aagcaagaac tccccgttca gcccgactgc
     7381 tgcgccttat ccggtaactg ttcacttgag tccaacccgg aaaagcacgg taaaacgcca
     7441 ctggcagcag ccattggtaa ctgggagttc gcagaagatt tgtttagcta aacacgcggt
     7501 tgctcttgaa gtgtgcgcca aagtccggct acactggaag gacagatttg gttgctgtgc
     7561 tctgcgaaag ccagttacca cggttaagca gttccccaac tgacttaacc ttcgatcaaa
     7621 ccacctcccc aggtggtttt ttcgtttaca gggcaaaaga ttacgcgcag aaaaaaagga
     7681 tctcaagaag atcctttgat cttttctact gaaccgctct agatttcagt gcaatttatc
     7741 tcttcaaatg tagcacctga agtcagcccc atacgatata agttgtaatt ctcatgttag
     7801 tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcgg
//
