LOCUS       sequence-362136-        7149 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of nuclease inactive SuperFi-Cas9 ABE8e base 
            editor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7149)
  AUTHORS   Kulcsar PI, Talas A, Ligeti Z, Krausz SL, Welker E
  TITLE     SuperFi-Cas9 exhibits remarkable fidelity but severely reduced 
            activity yet works effectively with ABE8e.
  JOURNAL   Nat Commun. 2022 Nov 11;13(1):6858. doi: 10.1038/s41467-022-34527-8.
  PUBMED    36369279
REFERENCE   2  (bases 1 to 7149)
  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: "2022-11-11"; volume: "13(1):6858. doi"; pages: " 
            10.1038/s41467-022-34527-8"
COMMENT     Sequence Label: pAT15545_dABE8e-SuperFi-Cas9
FEATURES             Location/Qualifiers
     source          1..7149
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        109..320
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             382..402
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(997..1020,1021..1023,1024..3579,3580..3582,3583..5097)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the N863A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVDGDDKVDDDDKMIAKSEDEIGDATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5098..5145
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(5173..5190)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5179..5386
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(5451..6039)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5531..5550)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6210..7001,7002..7070))
                     /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     6833..6852
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7071..7149,1..26))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtccag
       61 cagagatcca ctttggcgcc ggctcgagtg gctccggtgc ccgtcagtgg gcagagcgca
      121 catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacc ggtgcctaga
      181 gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg
      241 agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg
      301 ggtttgccgc cagaacacag gtgtcgtgac gcgggatccg ccaccatgaa acggacagcc
      361 gacggaagcg agttcgagtc accaaagaag aagcggaaag tctctgaggt ggagttttcc
      421 cacgagtact ggatgagaca tgccctgacc ctggccaaga gggcacggga tgagagggag
      481 gtgcctgtgg gagccgtgct ggtgctgaac aatagagtga tcggcgaggg ctggaacaga
      541 gccatcggcc tgcacgaccc aacagcccat gccgaaatta tggccctgag acagggcggc
      601 ctggtcatgc agaactacag actgattgac gccaccctgt acgtgacatt cgagccttgc
      661 gtgatgtgcg ccggcgccat gatccactct aggatcggcc gcgtggtgtt tggcgtgagg
      721 aactcaaaaa gaggcgccgc aggctccctg atgaacgtgc tgaactaccc cggcatgaat
      781 caccgcgtcg aaattaccga gggaatcctg gcagatgaat gtgccgccct gctgtgcgat
      841 ttctatcgga tgcctagaca ggtgttcaat gctcagaaga aggcccagag ctccatcaac
      901 tccggaggat ctagcggagg ctcctctggc tctgagacac ctggcacaag cgagagcgca
      961 acacctgaat cctcaggtgg tagctctggg gggtctgaca agaagtacag catcggcctg
     1021 gccatcggca ccaactctgt gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc
     1081 aagaaattca aggtgctggg caacaccgac cggcacagca tcaagaagaa cctgatcgga
     1141 gccctgctgt tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag aaccgccaga
     1201 agaagataca ccagacggaa gaaccggatc tgctatctgc aagagatctt cagcaacgag
     1261 atggccaagg tggacgacag cttcttccac agactggaag agtccttcct ggtggaagag
     1321 gataagaagc acgagcggca ccccatcttc ggcaacatcg tggacgaggt ggcctaccac
     1381 gagaagtacc ccaccatcta ccacctgaga aagaaactgg tggacagcac cgacaaggcc
     1441 gacctgcggc tgatctatct ggccctggcc cacatgatca agttccgggg ccacttcctg
     1501 atcgagggcg acctgaaccc cgacaacagc gacgtggaca agctgttcat ccagctggtg
     1561 cagacctaca accagctgtt cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag
     1621 gccatcctgt ctgccagact gagcaagagc agacggctgg aaaatctgat cgcccagctg
     1681 cccggcgaga agaagaatgg cctgttcggc aacctgattg ccctgagcct gggcctgacc
     1741 cccaacttca agagcaactt cgacctggcc gaggatgcca aactgcagct gagcaaggac
     1801 acctacgacg acgacctgga caacctgctg gcccagatcg gcgaccagta cgccgacctg
     1861 tttctggccg ccaagaacct gtccgacgcc atcctgctga gcgacatcct gagagtgaac
     1921 accgagatca ccaaggcccc cctgagcgcc tctatgatca agagatacga cgagcaccac
     1981 caggacctga ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa gtacaaagag
     2041 attttcttcg accagagcaa gaacggctac gccggctaca ttgacggcgg agccagccag
     2101 gaagagttct acaagttcat caagcccatc ctggaaaaga tggacggcac cgaggaactg
     2161 ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc ggaccttcga caacggcagc
     2221 atcccccacc agatccacct gggagagctg cacgccattc tgcggcggca ggaagatttt
     2281 tacccattcc tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc
     2341 tactacgtgg gccctctggc caggggaaac agcagattcg cctggatgac cagaaagagc
     2401 gaggaaacca tcaccccctg gaacttcgag gaagtggtgg acaagggcgc ttccgcccag
     2461 agcttcatcg agcggatgac caacttcgat aagaacctgc ccaacgagaa ggtgctgccc
     2521 aagcacagcc tgctgtacga gtacttcacc gtgtataacg agctgaccaa agtgaaatac
     2581 gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg
     2641 gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga ggactacttc
     2701 aagaaaatcg agtgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc
     2761 tccctgggca cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat
     2821 gaggaaaacg aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga
     2881 gagatgatcg aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag
     2941 cagctgaagc ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc
     3001 atccgggaca agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc
     3061 aacagaaact tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag
     3121 aaagcccagg tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc
     3181 agccccgcca ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa
     3241 gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc
     3301 acccagaagg gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa
     3361 gagctgggca gccagatcct gaaagaacac ccagtggaaa acacccagct gcagaacgag
     3421 aagctgtacc tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac
     3481 atcaaccggc tgtccgacta cgatgtggac gccatcgtgc ctcagagctt tctgaaggac
     3541 gactccatcg acaacaaggt gctgaccaga agcgacaaga accggggcaa gagcgacaac
     3601 gtgccctccg aagaggtcgt gaagaagatg aagaactact ggcggcagct gctgaacgcc
     3661 aagctgatta cccagagaaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc
     3721 gaactggata aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag
     3781 cacgtggcac agatcctgga ctcccggatg aacactaagt acgacgagaa tgacaagctg
     3841 atccgggaag tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat
     3901 ttccagtttt acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcgtacctg
     3961 aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg
     4021 gacggcgacg acaaggtgga cgacgacgac aagatgatcg ccaagagcga agacgaaatc
     4081 ggcgacgcta ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag
     4141 attaccctgg ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaacc
     4201 ggggagatcg tgtgggataa gggccgggat tttgccaccg tgcggaaagt gctgagcatg
     4261 ccccaagtga atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct
     4321 atcctgccca agaggaacag cgataagctg atcgccagaa agaaggactg ggaccctaag
     4381 aagtacggcg gcttcgacag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg
     4441 gaaaagggca agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg
     4501 gaaagaagca gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa
     4561 gtgaaaaagg acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc
     4621 cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc
     4681 tccaaatatg tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc
     4741 gaggataatg agcagaaaca gctgtttgtg gaacagcaca agcactacct ggacgagatc
     4801 atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa
     4861 gtgctgtccg cctacaacaa gcaccgggat aagcccatca gagagcaggc cgagaatatc
     4921 atccacctgt ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc
     4981 accatcgacc ggaagaggta caccagcacc aaagaggtgc tggacgccac cctgatccac
     5041 cagagcatca ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggcgacaag
     5101 cgtcctgctg ctactaagaa agctggtcaa gctaagaaaa agaaatgaca tgtcctagag
     5161 ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     5221 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     5281 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg
     5341 acagcaaggg ggaggattgg gaagagaata gcaggcatgc tggggagcgg ccgctgagca
     5401 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     5461 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     5521 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     5581 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     5641 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     5701 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5761 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5821 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5881 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5941 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     6001 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     6061 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     6121 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     6181 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     6241 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     6301 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     6361 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     6421 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     6481 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     6541 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     6601 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     6661 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6721 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6781 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6841 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6901 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6961 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     7021 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     7081 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     7141 tgtatttag
//
