LOCUS       sequence-335285-        7855 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses fusion protein pmCDA1-T7RNAP in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7855)
  AUTHORS   Alvarez B, Mencia M, de Lorenzo V, Fernandez LA
  TITLE     In vivo diversification of target genomic sites using processive 
            base deaminase fusions blocked by dCas9.
  JOURNAL   Nat Commun. 2020 Dec 22;11(1):6436. doi: 10.1038/s41467-020-20230-z.
  PUBMED    33353963
REFERENCE   2  (bases 1 to 7855)
  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."; date: "2020-12-22"; volume: "11(1):6436. doi"; pages: " 
            10.1038/s41467-020-20230-z"
COMMENT     Sequence Label: pSEVA221pmCDA1-T7RNAP
FEATURES             Location/Qualifiers
     source          1..7855
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      2300..2394
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(2515..2534)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2526..3341
                     /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"
     oriT            3508..3616
                     /note="incP origin of transfer"
     CDS             complement(3677..4825)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     IVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     rep_origin      5218..5749
                     /label=oriV
                     /note="incP origin of replication"
     terminator      complement(5880..5966)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(6009..6635)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSLRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     6540..6559
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        6651..6706
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    6657..6675
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    6687..6705
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             6770..7396
                     /codon_start=1
                     /product="cytidine deaminase from the sea lamprey 
                     Petromyzon marinus (Rogozin et al., 2007)"
                     /label=PmCDA1
                     /translation="MGTDAEYVRIHEKLDIYTFKKQFFNNKKSVSHRCYVLFELKRRGE
                     RRACFWGYAVNKPQSGTERGIHAEIFSIRKVEEYLRDNPGQFTINWYSSWSPCADCAEK
                     ILEWYNQELRGNGHTLKIWACKLYYEKNARNQIGLWNLRDNGVGLNVMVSEHYQCCRKI
                     FIQSSHNQLNENRWLEKTLKRAEKRRSELSIMIQVKILHTTKSPAV"
ORIGIN
        1 actctggctt gcctaaccag tgctgacaat acaaccgttc aggctgtagc aagcgcaatc
       61 ggtcgggcca ttgaggacga ggctcgcttc ggtcgtatcc gtgaccttga agctaagcac
      121 ttcaagaaaa acgttgagga acaactcaac aagcgcgtag ggcacgtcta caagaaagca
      181 tttatgcaag ttgtcgaggc tgacatgctc tctaagggtc tactcggtgg cgaggcgtgg
      241 tcttcgtggc ataaggaaga ctctattcat gtaggagtac gctgcatcga gatgctcatt
      301 gagtcaaccg gaatggttag cttacaccgc caaaatgctg gcgtagtagg tcaagactct
      361 gagactatcg aactcgcacc tgaatacgct gaggctatcg caacccgtgc aggtgcgctg
      421 gctggcatct ctccgatgtt ccaaccttgc gtagttcctc ctaagccgtg gactggcatt
      481 actggtggtg gctattgggc taacggtcgt cgtcctctgg cgctggtgcg tactcacagt
      541 aagaaagcac tgatgcgcta cgaagacgtt tacatgcctg aggtgtacaa agcgattaac
      601 attgcgcaaa acaccgcatg gaaaatcaac aagaaagtcc tagcggtcgc caacgtaatc
      661 accaagtgga agcattgtcc ggtcgaggac atccctgcga ttgagcgtga agaactcccg
      721 atgaaaccgg aagacatcga catgaatcct gaggctctca ccgcgtggaa acgtgctgcc
      781 gctgctgtgt accgcaagga caaggctcgc aagtctcgcc gtatcagcct tgagttcatg
      841 cttgagcaag ccaataagtt tgctaaccat aaggccatct ggttccctta caacatggac
      901 tggcgcggtc gtgtttacgc tgtgtcaatg ttcaacccgc aaggtaacga tatgaccaaa
      961 ggactgctta cgctggcgaa aggtaaacca atcggtaagg aaggttacta ctggctgaaa
     1021 atccacggtg caaactgtgc gggtgtcgat aaggttccgt tccctgagcg catcaagttc
     1081 attgaggaaa accacgagaa catcatggct tgcgctaagt ctccactgga gaacacttgg
     1141 tgggctgagc aagattctcc gttctgcttc cttgcgttct gctttgagta cgctggggta
     1201 cagcaccacg gcctgagcta taactgctcc cttccgctgg cgtttgacgg gtcttgctct
     1261 ggcatccagc acttctccgc gatgctccga gatgaggtag gtggtcgcgc ggttaacttg
     1321 cttcctagtg aaaccgttca ggacatctac gggattgttg ctaagaaagt caacgagatt
     1381 ctacaagcag acgcaatcaa tgggaccgat aacgaagtag ttaccgtgac cgatgagaac
     1441 actggtgaaa tctctgagaa agtcaagctg ggcactaagg cactggctgg tcaatggctg
     1501 gcttacggtg ttactcgcag tgtgactaag cgttcagtca tgacgctggc ttacgggtcc
     1561 aaagagttcg gcttccgtca acaagtgctg gaagatacca ttcagccagc tattgattcc
     1621 ggcaagggtc tgatgttcac tcagccgaat caggctgctg gatacatggc taagctgatt
     1681 tgggaatctg tgagcgtgac ggtggtagct gcggttgaag caatgaactg gcttaagtct
     1741 gctgctaagc tgctggctgc tgaggtcaaa gataagaaga ctggagagat tcttcgcaag
     1801 cgttgcgctg tgcattgggt aactcctgat ggtttccctg tgtggcagga atacaagaag
     1861 cctattcaga cgcgcttgaa cctgatgttc ctcggtcagt tccgcttaca gcctaccatt
     1921 aacaccaaca aagatagcga gattgatgca cacaaacagg agtctggtat cgctcctaac
     1981 tttgtacaca gccaagacgg tagccacctt cgtaagactg tagtgtgggc acacgagaag
     2041 tacggaatcg aatcttttgc actgattcac gactccttcg gtaccattcc ggctgacgct
     2101 gcgaacctgt tcaaagcagt gcgcgaaact atggttgaca catatgagtc ttgtgatgta
     2161 ctggctgatt tctacgacca gttcgctgac cagttgcacg agtctcaatt ggacaaaatg
     2221 ccagcacttc cggctaaagg taacttgaac ctccgtgaca tcttagagtc ggacttcgcg
     2281 ttcgcgtaaa ctagtcttgg actcctgttg atagatccag taatgacctc agaactccat
     2341 ctggatttgt tcagaacgct cggttgccgc cgggcgtttt ttattggtga gaatccaggg
     2401 gtccccaata attacgattt aaatttgtgt ctcaaaatct ctgatgttac attgcacaag
     2461 ataaaaatat atcatcatga acaataaaac tgtctgctta cataaacagt aatacaaggg
     2521 gtgttatgag ccatattcag cgtgaaacga gctgtagccg tccgcgtctg aacagcaaca
     2581 tggatgcgga tctgtatggc tataaatggg cgcgtgataa cgtgggtcag agcggcgcga
     2641 ccatttatcg tctgtatggc aaaccggatg cgccggaact gtttctgaaa catggcaaag
     2701 gcagcgtggc gaacgatgtg accgatgaaa tggtgcgtct gaactggctg accgaattta
     2761 tgccgctgcc gaccattaaa cattttattc gcaccccgga tgatgcgtgg ctgctgacca
     2821 ccgcgattcc gggcaaaacc gcgtttcagg tgctggaaga atatccggat agcggcgaaa
     2881 acattgtgga tgcgctggcc gtgtttctgc gtcgtctgca tagcattccg gtgtgcaact
     2941 gcccgtttaa cagcgatcgt gtgtttcgtc tggcccaggc gcagagccgt atgaacaacg
     3001 gcctggtgga tgcgagcgat tttgatgatg aacgtaacgg ctggccggtg gaacaggtgt
     3061 ggaaagaaat gcataaactg ctgccgttta gcccggatag cgtggtgacc cacggcgatt
     3121 ttagcctgga taacctgatt ttcgatgaag gcaaactgat tggctgcatt gatgtgggcc
     3181 gtgtgggcat tgcggatcgt tatcaggatc tggccattct gtggaactgc ctgggcgaat
     3241 ttagcccgag cctgcaaaaa cgtctgtttc agaaatatgg cattgataat ccggatatga
     3301 acaaactgca atttcatctg atgctggatg aatttttcta ataattaatt ggaccgcggt
     3361 ccgcgcgttg tccttttccg ctgcataacc ctgcttcggg gtcattatag cgattttttc
     3421 ggtatatcca tcctttttcg cacgatatac aggattttgc caaagggttc gtgtagactt
     3481 tccttggtgt atccaacggc gtcagccggg caggataggt gaagtaggcc cacccgcgag
     3541 cgggtgttcc ttcttcactg tcccttattc gcacctggcg gtgctcaacg ggaatcctgc
     3601 tctgcgaggc tggccgtagg ccggccgcga tgcaggtggc tgctgaaccc ccagccggaa
     3661 ctgaccccac aaggccctag cgtttgcaat gcaccaggtc atcattgacc caggcgtgtt
     3721 ccaccaggcc gctgcctcgc aactcttcgc aggcttcgcc gacctgctcg cgccacttct
     3781 tcacgcgggt ggaatccgat ccgcacatga ggcggaaggt ttccagcttg agcgggtacg
     3841 gctcccggtg cgagctgaaa tagtcgaaca tccgtcgggc cgtcggcgac agcttgcggt
     3901 acttctccca tatgaatttc gtgtagtggt cgccagcaaa cagcacgacg atttcctcgt
     3961 cgatcaggac ctggcaacgg gacgttttct tgccacggtc caggacgcgg aagcggtgca
     4021 gcagcgacac cgattccagg tgcccaacgc ggtcggacgt gaagcccatc gccgtcgcct
     4081 gtaggcgcga caggcattcc tcggccttcg tgtaataccg gccattgatc gaccagccca
     4141 ggtcctggca aagctcgtag aacgtgaagg tgatcggctc gccgataggg gtgcgcttcg
     4201 cgtactccaa cacctgctgc cacaccagtt cgtcatcgtc ggcccgcagc tcgacgccgg
     4261 tgtaggtgat cttcacgtcc ttgttgacgt ggaaaatgac cttgttttgc agcgcctcgc
     4321 gcgggatttt cttgttgcgc gtggtgaaca gggcagagcg ggccgtgtcg tttggcatcg
     4381 ctcgcatcgt gtccggccac ggcgcaatat cgaacaagga aagctgcatt tccttgatct
     4441 gctgcttcgt gtgtttcagc aacgcggcct gcttggcttc gctgacctgt tttgccaggt
     4501 cctcgccggc ggtttttcgc ttcttggtcg tcatagttcc tcgcgtgtcg atggtcatcg
     4561 acttcgccaa acctgccgcc tcctgttcga gacgacgcga acgctccacg gcggccgatg
     4621 gcgcgggcag ggcaggggga gccagttgca cgctgtcgcg ctcgatcttg gccgtagctt
     4681 gctggactat cgagccgacg gactggaagg tttcgcgggg cgcacgcatg acggtgcggc
     4741 ttgcgatggt ttcggcatcc tcggcggaaa accccgcgtc gatcagttct tgcctgtatg
     4801 ccttccggtc aaacgtccga ttcattcacc ctccttgcgg gattgccccg gaattaattc
     4861 cccggatcga tccgtcgatc ttgatcccct gcgccatcag atccttggcg gcaagaaagc
     4921 catccagttt actttgcagg gcttcccaac cttaccagag ggcgccccag ctggcaattc
     4981 cggttcgctt gctgtccata aaaccgccca gtctagctat cgccatgtaa gcccactgca
     5041 agctacctgc tttctctttg cgcttgcgtt ttcccttgtc cagatagccc agtagctgac
     5101 attcatccgg ggtcagcacc gtttctgcgg actggctttc tacgtggctg ccatttttgg
     5161 ggtgaggccg ttcgcggccg aggggcgcag cccctggggg gatgggaggc ccgcgttagc
     5221 gggccgggag ggttcgagaa gggggggcac cccccttcgg cgtgcgcggt cacgcgcaca
     5281 gggcgcagcc ctggttaaaa acaaggttta taaatattgg tttaaaagca ggttaaaaga
     5341 caggttagcg gtggccgaaa aacgggcgga aacccttgca aatgctggat tttctgcctg
     5401 tggacagccc ctcaaatgtc aataggtgcg cccctcatct gtcagcactc tgcccctcaa
     5461 gtgtcaagga tcgcgcccct catctgtcag tagtcgcgcc cctcaagtgt caataccgca
     5521 gggcacttat ccccaggctt gtccacatca tctgtgggaa actcgcgtaa aatcaggcgt
     5581 tttcgccgat ttgcgaggct ggccagctcc acgtcgccgg ccgaaatcga gcctgcccct
     5641 catctgtcaa cgccgcgccg ggtgagtcgg cccctcaagt gtcaacgtcc gcccctcatc
     5701 tgtcagtgag ggccaagttt tccgcgaggt atccacaacg ccggcggccc tacatggctc
     5761 tgctgtagtg agtgggttgc gctccggcag cggtcctgat cccccgcaga aaaaaaggat
     5821 ctcaagaaga tcctttgatc ttttctacgg cgcgccgagc tcgaattcta gggcggcgga
     5881 tttgtcctac tcaggagagc gttcaccgac aaacaacaga taaaacgaaa ggcccagtct
     5941 ttcgactgag cctttcgttt tatttgatgc ctctagcacg cctgccagga attggggatc
     6001 ggagatcttt aagacccact ttcacattta agttgttttt ctaatccgca tataatcaat
     6061 tcaaggccga ataagaaggc tggctctgca ccttggtgat caaataattc gatagcttgt
     6121 cgtaataatg gcggcatact atcagtagta ggtgtttccc tttcttcttt agcgacttga
     6181 tgctcttgat cttccaatac gcaacctaaa gtaaaatgcc ccacagcgct gagtgcatat
     6241 aatgcattct ctagtgaaaa accttgttgg cataaaaagg ctaattgatt ttcgagagtt
     6301 tcatactgtt tttctgtagg ccgtgtacct aaatgtactt ttgctccatc gcgatgactt
     6361 agtaaagcac atctaagact tttagcgtta ttacgtaaaa aatcttgcca gctttcccct
     6421 tctaaagggc aaaagtgagt atggtgccta tctaacatct caatggctaa ggcgtcgagc
     6481 aaagcccgct tattttttac atgccaatac aatgtaggct gctctacacc tagcttctgg
     6541 gcgagtttac gggttgttaa accttcgatt ccgacctcat taagcagctc taatgcgctg
     6601 ttaatcactt tacttttatc taatctggac atcattaatt cctaattttt gttgacactc
     6661 tatcattgat agagttattt taccactccc tatcagtgat agagaaaagt gaatctagag
     6721 tcgacaagaa ggagatatac atatggcaag catgaccggt ggtcagcaga tgggcaccga
     6781 tgcagaatat gttcgtattc atgaaaagct ggacatctat accttcaaaa aacagttctt
     6841 caacaacaaa aagagcgtga gccatcgttg ctatgttctg tttgaactga aacgtcgtgg
     6901 tgaacgtcgt gcatgttttt ggggttatgc agttaataaa ccgcagagcg gcaccgaacg
     6961 tggtattcat gcagaaatct ttagcattcg caaagtggaa gaatatctgc gtgataatcc
     7021 gggtcagttt accattaatt ggtatagcag ctggtcaccg tgtgcagatt gtgcagaaaa
     7081 aattctggaa tggtataacc aagaactgcg tggtaatggt cataccctga aaatttgggc
     7141 atgtaaactg tactacgaaa agaatgcccg taatcagatt ggtctgtgga atctgcgcga
     7201 taacggtgtt ggtctgaatg ttatggttag cgaacattat cagtgctgcc gcaaaatctt
     7261 tattcagagc agccataatc agctgaatga aaatcgttgg ctggaaaaaa ccctgaaacg
     7321 tgccgaaaaa cgtcgtagcg aactgagcat tatgattcag gttaaaattc tgcacaccac
     7381 caaaagtccg gcagttctgc agggtggtgg tagcggtggt ggttcaggtg gtggcagtgg
     7441 cggtggaagt ggtggcggtt ctggcggagg atcaggtggc ggtagcaagc ttatgaacac
     7501 gattaacatc gctaagaacg acttctctga catcgaactg gctgctatcc cgttcaacac
     7561 tctggctgac cattacggtg agcgtttagc tcgcgaacag ttggcccttg agcatgagtc
     7621 ttacgagatg ggtgaagcac gcttccgcaa gatgtttgag cgtcaactta aagctggtga
     7681 ggttgcggat aacgctgccg ccaagcctct catcactacc ctactcccta agatgattgc
     7741 acgcatcaac gactggtttg aggaagtgaa agctaagcgc ggcaagcgcc cgacagcctt
     7801 ccagttcctg caagaaatca agccggaagc cgtagcgtac atcaccatta agacc
//
