LOCUS       sequence-362137-        7149 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of 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: pAT15546_nABE8e-SuperFi-Cas9
FEATURES             Location/Qualifiers
     source          1..7149
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        129..340
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             402..422
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1017..5117
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVDGDDKVDDDDKMIAKSEDEIGDATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5118..5165
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(5193..5210)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5199..5406
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(5471..6059)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5551..5570)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6230..7021,7022..7090))
                     /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     6853..6872
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7091..7149,1..46))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
       61 aagtgccacc tgacgtccag cagagatcca ctttggcgcc ggctcgagtg gctccggtgc
      121 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
      181 caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
      241 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
      301 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtgtcgtgac gcgggatccg
      361 ccaccatgaa acggacagcc gacggaagcg agttcgagtc accaaagaag aagcggaaag
      421 tctctgaggt ggagttttcc cacgagtact ggatgagaca tgccctgacc ctggccaaga
      481 gggcacggga tgagagggag gtgcctgtgg gagccgtgct ggtgctgaac aatagagtga
      541 tcggcgaggg ctggaacaga gccatcggcc tgcacgaccc aacagcccat gccgaaatta
      601 tggccctgag acagggcggc ctggtcatgc agaactacag actgattgac gccaccctgt
      661 acgtgacatt cgagccttgc gtgatgtgcg ccggcgccat gatccactct aggatcggcc
      721 gcgtggtgtt tggcgtgagg aactcaaaaa gaggcgccgc aggctccctg atgaacgtgc
      781 tgaactaccc cggcatgaat caccgcgtcg aaattaccga gggaatcctg gcagatgaat
      841 gtgccgccct gctgtgcgat ttctatcgga tgcctagaca ggtgttcaat gctcagaaga
      901 aggcccagag ctccatcaac tccggaggat ctagcggagg ctcctctggc tctgagacac
      961 ctggcacaag cgagagcgca acacctgaat cctcaggtgg tagctctggg gggtctgaca
     1021 agaagtacag catcggcctg gccatcggca ccaactctgt gggctgggcc gtgatcaccg
     1081 acgagtacaa ggtgcccagc aagaaattca aggtgctggg caacaccgac cggcacagca
     1141 tcaagaagaa cctgatcgga gccctgctgt tcgacagcgg cgaaacagcc gaggccaccc
     1201 ggctgaagag aaccgccaga agaagataca ccagacggaa gaaccggatc tgctatctgc
     1261 aagagatctt cagcaacgag atggccaagg tggacgacag cttcttccac agactggaag
     1321 agtccttcct ggtggaagag gataagaagc acgagcggca ccccatcttc ggcaacatcg
     1381 tggacgaggt ggcctaccac gagaagtacc ccaccatcta ccacctgaga aagaaactgg
     1441 tggacagcac cgacaaggcc gacctgcggc tgatctatct ggccctggcc cacatgatca
     1501 agttccgggg ccacttcctg atcgagggcg acctgaaccc cgacaacagc gacgtggaca
     1561 agctgttcat ccagctggtg cagacctaca accagctgtt cgaggaaaac cccatcaacg
     1621 ccagcggcgt ggacgccaag gccatcctgt ctgccagact gagcaagagc agacggctgg
     1681 aaaatctgat cgcccagctg cccggcgaga agaagaatgg cctgttcggc aacctgattg
     1741 ccctgagcct gggcctgacc cccaacttca agagcaactt cgacctggcc gaggatgcca
     1801 aactgcagct gagcaaggac acctacgacg acgacctgga caacctgctg gcccagatcg
     1861 gcgaccagta cgccgacctg tttctggccg ccaagaacct gtccgacgcc atcctgctga
     1921 gcgacatcct gagagtgaac accgagatca ccaaggcccc cctgagcgcc tctatgatca
     1981 agagatacga cgagcaccac caggacctga ccctgctgaa agctctcgtg cggcagcagc
     2041 tgcctgagaa gtacaaagag attttcttcg accagagcaa gaacggctac gccggctaca
     2101 ttgacggcgg agccagccag gaagagttct acaagttcat caagcccatc ctggaaaaga
     2161 tggacggcac cgaggaactg ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc
     2221 ggaccttcga caacggcagc atcccccacc agatccacct gggagagctg cacgccattc
     2281 tgcggcggca ggaagatttt tacccattcc tgaaggacaa ccgggaaaag atcgagaaga
     2341 tcctgacctt ccgcatcccc tactacgtgg gccctctggc caggggaaac agcagattcg
     2401 cctggatgac cagaaagagc gaggaaacca tcaccccctg gaacttcgag gaagtggtgg
     2461 acaagggcgc ttccgcccag agcttcatcg agcggatgac caacttcgat aagaacctgc
     2521 ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga gtacttcacc gtgtataacg
     2581 agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg
     2641 agcagaaaaa ggccatcgtg gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc
     2701 agctgaaaga ggactacttc aagaaaatcg agtgcttcga ctccgtggaa atctccggcg
     2761 tggaagatcg gttcaacgcc tccctgggca cataccacga tctgctgaaa attatcaagg
     2821 acaaggactt cctggacaat gaggaaaacg aggacattct ggaagatatc gtgctgaccc
     2881 tgacactgtt tgaggacaga gagatgatcg aggaacggct gaaaacctat gcccacctgt
     2941 tcgacgacaa agtgatgaag cagctgaagc ggcggagata caccggctgg ggcaggctga
     3001 gccggaagct gatcaacggc atccgggaca agcagtccgg caagacaatc ctggatttcc
     3061 tgaagtccga cggcttcgcc aacagaaact tcatgcagct gatccacgac gacagcctga
     3121 cctttaaaga ggacatccag aaagcccagg tgtccggcca gggcgatagc ctgcacgagc
     3181 acattgccaa tctggccggc agccccgcca ttaagaaggg catcctgcag acagtgaagg
     3241 tggtggacga gctcgtgaaa gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa
     3301 tggccagaga gaaccagacc acccagaagg gacagaagaa cagccgcgag agaatgaagc
     3361 ggatcgaaga gggcatcaaa gagctgggca gccagatcct gaaagaacac cccgtggaaa
     3421 acacccagct gcagaacgag aagctgtacc tgtactacct gcagaatggg cgggatatgt
     3481 acgtggacca ggaactggac atcaaccggc tgtccgacta cgatgtggac catatcgtgc
     3541 ctcagagctt tctgaaggac gactccatcg acaacaaggt gctgaccaga agcgacaaga
     3601 accggggcaa gagcgacaac gtgccctccg aagaggtcgt gaagaagatg aagaactact
     3661 ggcggcagct gctgaacgcc aagctgatta cccagagaaa gttcgacaat ctgaccaagg
     3721 ccgagagagg cggcctgagc gaactggata aggccggctt catcaagaga cagctggtgg
     3781 aaacccggca gatcacaaag cacgtggcac agatcctgga ctcccggatg aacactaagt
     3841 acgacgagaa tgacaagctg atccgggaag tgaaagtgat caccctgaag tccaagctgg
     3901 tgtccgattt ccggaaggat ttccagtttt acaaagtgcg cgagatcaac aactaccacc
     3961 acgcccacga cgcgtacctg aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta
     4021 agctggaaag cgagttcgtg gacggcgacg acaaggtgga cgacgacgac aagatgatcg
     4081 ccaagagcga agacgaaatc ggcgacgcta ccgccaagta cttcttctac agcaacatca
     4141 tgaacttttt caagaccgag attaccctgg ccaacggcga gatccggaag cggcctctga
     4201 tcgagacaaa cggcgaaacc ggggagatcg tgtgggataa gggccgggat tttgccaccg
     4261 tgcggaaagt gctgagcatg ccccaagtga atatcgtgaa aaagaccgag gtgcagacag
     4321 gcggcttcag caaagagtct atcctgccca agaggaacag cgataagctg atcgccagaa
     4381 agaaggactg ggaccctaag aagtacggcg gcttcgacag ccccaccgtg gcctattctg
     4441 tgctggtggt ggccaaagtg gaaaagggca agtccaagaa actgaagagt gtgaaagagc
     4501 tgctggggat caccatcatg gaaagaagca gcttcgagaa gaatcccatc gactttctgg
     4561 aagccaaggg ctacaaagaa gtgaaaaagg acctgatcat caagctgcct aagtactccc
     4621 tgttcgagct ggaaaacggc cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg
     4681 gaaacgaact ggccctgccc tccaaatatg tgaacttcct gtacctggcc agccactatg
     4741 agaagctgaa gggctccccc gaggataatg agcagaaaca gctgtttgtg gaacagcaca
     4801 agcactacct ggacgagatc atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg
     4861 ccgacgctaa tctggacaaa gtgctgtccg cctacaacaa gcaccgggat aagcccatca
     4921 gagagcaggc cgagaatatc atccacctgt ttaccctgac caatctggga gcccctgccg
     4981 ccttcaagta ctttgacacc accatcgacc ggaagaggta caccagcacc aaagaggtgc
     5041 tggacgccac cctgatccac cagagcatca ccggcctgta cgagacacgg atcgacctgt
     5101 ctcagctggg aggcgacaag cgtcctgctg ctactaagaa agctggtcaa gctaagaaaa
     5161 agaaatgaca tgtcctagag ctcgctgatc agcctcgact gtgccttcta gttgccagcc
     5221 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     5281 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     5341 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagagaata gcaggcatgc
     5401 tggggagcgg ccgctgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5461 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5521 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     5581 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5641 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     5701 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     5761 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     5821 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5881 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     5941 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     6001 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     6061 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6121 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     6181 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     6241 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     6301 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     6361 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     6421 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     6481 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     6541 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     6601 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     6661 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     6721 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     6781 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     6841 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     6901 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     6961 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     7021 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     7081 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     7141 tgagcggat
//
