LOCUS       sequence-432902-        5652 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Unfused click editor construct expressing PCV2-ePhi29(D169A), 
            expressed from CMV or T7 promoters..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5652)
  AUTHORS   Ferreira da Silva J, Tou CJ, King EM, Eller ML, Rufino-Ramos D, Ma 
            L, Cromwell CR, Metovic J, Benning FMC, Chao LH, Eichler FS, 
            Kleinstiver BP
  TITLE     Click editing enables programmable genome writing using DNA 
            polymerases and HUH endonucleases.
  JOURNAL   Nat Biotechnol. 2024 Jul 22. doi: 10.1038/s41587-024-02324-x.
  PUBMED    39039307
REFERENCE   2  (bases 1 to 5652)
  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 
            Biotechnol. 2024 Jul 22. doi: 10.1038/s41587-024-02324-x."
COMMENT     Sequence Label: Unfused click editor construct; PCV2-ePhi29 fu...
FEATURES             Location/Qualifiers
     source          1..5652
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(171..962,963..1031))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     794..813
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1032..1136)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1215..1234)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1406..1785
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1786..1989
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1940..1960
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2031..2050
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2031..2049
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4292..4312
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4321..4338
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4361..4378)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4367..4591
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4662..4678)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4662..4678)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4675..4697)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4686..4702
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4710..4716,4717..4734,4735..4740))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4755..4776
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4893..4910)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5064..5652)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5144..5163)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
       61 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
      121 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
      181 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
      241 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
      301 atgataccgc gcgaaccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
      361 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
      421 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
      481 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
      541 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
      601 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
      661 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
      721 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
      781 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
      841 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      901 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      961 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     1021 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     1081 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     1141 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atcgatctcc cgatccccta
     1201 gggtcgactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
     1261 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa
     1321 ggcttgaccg acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc
     1381 gatgtacggg ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca
     1441 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
     1501 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
     1561 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
     1621 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     1681 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     1741 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
     1801 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
     1861 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
     1921 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
     1981 agcagagctg gtttagtgaa ccgtcagatc cgctagagat ccgcggccgc taatacgact
     2041 cactataggg agagccgcca ccatgagccc atctaagaaa aacggcagaa gcggacctca
     2101 acctcacaag cggtgggtct ttaccctgaa caaccccagc gaagatgaga gaaagaagat
     2161 ccgggacctg cctatcagcc tgttcgacta cttcatcgtg ggcgaggaag gaaatgagga
     2221 aggccggacc ccccacctgc agggcttcgc caacttcgtg aaaaagcaga ccttcaacaa
     2281 ggtgaagtgg tacctgggcg ctagatgcca catcgagaag gccaagggca ccgaccagca
     2341 gaacaaagag tactgcagca aggaaggcaa tctgctgatg gaatgtggcg cccctagatc
     2401 ccagggacag agatctggag gatctagcgg aggatcctct ggcagcgaga caccaggaac
     2461 aagcgagtca gcaacaccag agagcagtgg cggcagcagc ggcggcagca gcaagcacat
     2521 gcctagaaag agatacagct gcgacttcga gacaacaacc aaggtcgagg actgcagagt
     2581 gtgggcttac ggatatatga atatcgagga ccactctgag tacaagatcg gaaatagcct
     2641 ggatgagttt atggcctggg ccctgaaagt gcaggccgac ctgtacttcc acaacctgaa
     2701 attcgacggc gccttcatca ttaactggct ggaacggaac ggctttaagt ggtccgccga
     2761 tggactgcca aacacctaca acaccatcat cagcagaacc ggccagtggt acatgatcga
     2821 catctgcctg ggctacaagg gcaaaagaaa gatccatacc gtgatctacg acagcctgaa
     2881 gaaactgcct ttccccgtga agaagatcgc taaagacttc aagctgacag tgctgaaggg
     2941 cgacattgat taccacaagg aaagacctgt gggatacaag atcacacctg aggaatatgc
     3001 ctacatcaag aacgccatcc agatcatcgc cgaggccctg ctgatccagt tcaagcaggg
     3061 cctggaccgg atgaccgctg gcagcgacag cctcaaagac ttcaaggaca ttatcaccac
     3121 taagaagttc aaaaaggtgt tcccaacact gtctcttgga ctggacaaga aggtgcggta
     3181 cgcctataga ggcggcttta cctggctgaa cgacagattc aaggaaaagg aaatcggcga
     3241 gggcatggtg tttgacgtga actccctcta ccctgcccag atgtacagcc ggctgctgcc
     3301 ctatggcgag cctatcgtgt ttgagggcaa gtacgtgtgg gatgaagatt accccctgca
     3361 catccagcac atccggtgcg agttcgagct gaaggagggc tacatcccaa caatccaaat
     3421 caaacggagc agattctaca agggcaacga gtacctgaaa agcagcggcg gcgaaatcgc
     3481 cgatctgtgg ctgagcaatg tggacctgga actgatgaag gaacactacg acctgtacaa
     3541 cgtggaatac atcagcggcc tgaagtttaa ggccacaacc ggcctgttca aagacttcat
     3601 cgataagtgg acctacatca aaacaaccag cgagggcgct atcaagcagc tggccaagct
     3661 gatgctgaat tctctgtacg gcaagttcgc cagcaaccct gatgtcacag gcaaggtgcc
     3721 ctacctgaag gaaaacggcg ccctgggctt cagactgggc gaggaagaaa ccaaggaccc
     3781 tgtgtacacc cctatgggtg ttttcatcac cgcctgggcc agatacacca ccatcaccgc
     3841 cgctcaggct tgttacgata gaatcatcta ttgtgatacc gactctatcc acctgaccgg
     3901 gaccgagatc cccgacgtga tcaaggatat tgtggacccc aagaaactgg gctactgggc
     3961 ccacgagtcc accttcaagc gggccaagta cctgagaccc aagacctaca tccaggacat
     4021 ctacatgaaa gaagtggacg gcgagctcgt ggaaggcagc cccgacgact acacagacat
     4081 caagctgagc gtgaagtgcg ccggcatgac cgataagatc aaaaaggagg tgacatttga
     4141 gaacttcaag gtgggattca gccgcaagat gaaacctaag cctgttcaag tgcctggcgg
     4201 tgttgtgctg gtggatgaca ccttcacaat caagtccggc ggaagctctg gtggcagcaa
     4261 gcggaccgcc gacggctctg aattcgagag ccctaagaag aaaagaaagg tgtgaccggt
     4321 catcatcacc atcaccattg agtttaaacc cgctgatcag cctcgactgt gccttctagt
     4381 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     4441 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     4501 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     4561 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc
     4621 tcgataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg
     4681 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     4741 gcctagggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     4801 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     4861 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     4921 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     4981 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5041 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     5101 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     5161 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     5221 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     5281 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     5341 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     5401 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     5461 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     5521 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     5581 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     5641 aaaggatctc aa
//
