LOCUS       sequence-384720-        6204 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses human codon-optimized SpCas9-NRCH-CGBE1 and blasticidin 
            resistance: EFS promoter-SpCas9-NRCH-CGBE1-NLS-FLAG-P2A-BSD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 6204)
  AUTHORS   Kim N, Choi S, Kim S, Song M, Seo JH, Min S, Park J, Cho SR, Kim HH
  TITLE     Deep learning models to predict the editing efficiencies and 
            outcomes of diverse base editors.
  JOURNAL   Nat Biotechnol. 2024 Mar;42(3):484-497. doi: 
            10.1038/s41587-023-01792-x
  PUBMED    37188916
REFERENCE   2  (bases 1 to 6204)
  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"; doi: "10"; journalName: 
            "Nat Biotechnol"; date: "2024-03"; volume: "42"; issue: "3"; pages: 
            "484-497"
COMMENT     Sequence Label: SpCas9-NRCH-CGBE1
FEATURES             Location/Qualifiers
     source          1..6204
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             37..57
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1552..5652
                     /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
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMVKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGIIPHQIHLGELHAILRRQGDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRLRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGGH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKGNSDKLIARKKDWDPKKYGGFNSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGVLQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTINRKQYNTTKEVLD
                     ATLIRQSITGLYETRIDLSQLGGD"
     CDS             5695..5715
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5716..5739
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             5749..5805
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             5806..6204
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
ORIGIN
        1 atgaaacgga cagccgacgg aagcgagttc gagtcaccaa agaagaagcg gaaagtcgcc
       61 aacgaactga cctggcacga cgtgctggcc gaagagaagc agcagcccta tttccttaac
      121 accctgcaga ccgtggccag cgagcggcag agcggcgtga ccatctaccc cccacagaaa
      181 gacgtgttta acgccttccg cttcacagag ctgggcgacg tgaaggtggt gatcctgggc
      241 caggacccct accacggccc cggccaggcc catggtctgg ccttcagcgt gcggcccggc
      301 atcgccatcc cccccagcct gctgaacatg tataaggagc tggaaaacac catccccggc
      361 ttcacccggc ccaatcacgg ctacctggag agctgggcgc ggcagggcgt gctgctgctc
      421 aacaccgtgc tgacggtacg ggccggccag gcgcatagcc acgccagcct gggctgggag
      481 accttcaccg ataaggtgat cagcctgatc aaccagcacc gggagggcgt ggtgtttctg
      541 ctgtggggca gccatgccca gaagaaaggc gccatcatcg acaagcagcg gcatcatgtg
      601 ctgaaagccc cccaccccag cccccttagc gcccaccggg gcttcttcgg ctgcaaccat
      661 ttcgtgctgg ccaatcagtg gctggaacaa cggggcgaga cccccattga ctggatgccc
      721 gtgttgcccg ccgagagcga gagcggcggg agcggcggga gcggggggag ctcctcagag
      781 actgggcctg tcgccgtcga tccaaccctg cgccgccgga ttgaacctca cgagtttgaa
      841 gtgttctttg acccccggga gctggccaag gagacatgcc tgctgtacga gatcaactgg
      901 ggaggcaggc actccatctg gaggcacacc tctcagaaca caaataagca cgtggaggtg
      961 aacttcatcg agaagtttac cacagagcgg tacttctgcc ccaataccag atgtagcatc
     1021 acatggtttc tgagctggtc cccttgcgga gagtgtagca gggccatcac cgagttcctg
     1081 tccagatatc cacacgtgac actgtttatc tacatcgcca ggctgtatca ccacgcagac
     1141 ccaaggaata ggcagggcct gcgcgatctg atcagctccg gcgtgaccat ccagatcatg
     1201 acagagcagg agtccggcta ctgctggcgg aacttcgtga attattctcc tagcaacgag
     1261 gcccactggc ctaggtaccc acacctgtgg gtgcgcctgt acgtgctgga gctgtattgc
     1321 atcatcctgg gcctgccccc ttgtctgaat atcctgcgga gaaagcagcc ccagctgacc
     1381 ttctttacaa tcgccctgca gtcttgtcac tatcagaggc tgccacccca catcctgtgg
     1441 gccacaggcc tgaagtctgg aggatctagc ggaggatcct ctggcagcga gacaccagga
     1501 acaagcgagt cagcaacacc agagagcagt ggcggcagca gcggcggcag cgacaagaag
     1561 tacagcatcg gcctggccat cggcaccaac tctgtgggct gggccgtgat caccgacgag
     1621 tacaaggtgc ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca cagcatcaag
     1681 aagaacctga tcggagccct gctgttcgac agcggcgaaa cagccgaggc cacccggctg
     1741 aagagaaccg ccagaagaag atacaccaga cggaagaacc ggatctgcta tctgcaagag
     1801 atcttcagca acgagatggc caaggtggac gacagcttct tccacagact ggaagagtcc
     1861 ttcctggtgg aagaggataa gaagcacgag cggcacccca tcttcggcaa catcgtggac
     1921 gaggtggcct accacgagaa gtaccccacc atctaccacc tgagaaagaa actggtggac
     1981 agcaccgaca aggccgacct gcggctgatc tatctggccc tggcccacat gatcaagttc
     2041 cggggccact tcctgatcga gggcgacctg aaccccgaca acagcgacgt ggacaagctg
     2101 ttcatccagc tggtgcagac ctacaaccag ctgttcgagg aaaaccccat caacgccagc
     2161 ggcgtggacg ccaaggccat cctgtctgcc agactgagca agagcagacg gctggaaaat
     2221 ctgatcgccc agctgcccgg cgagaagaag aatggcctgt tcggaaacct gattgccctg
     2281 agcctgggcc tgacccccaa cttcaagagc aacttcgacc tggccgagga tgccaaactg
     2341 cagctgagca aggacaccta cgacgacgac ctggacaacc tgctggccca gatcggcgac
     2401 cagtacgccg acctgtttct ggccgccaag aacctgtccg acgccatcct gctgagcgac
     2461 atcctgagag tgaacaccga gatcaccaag gcccccctga gcgcctctat ggtgaagaga
     2521 tacgacgagc accaccagga cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct
     2581 gagaagtaca aagagatttt cttcgaccag agcaagaacg gctacgccgg ctacattgac
     2641 ggcggagcca gccaggaaga gttctacaag ttcatcaagc ccatcctgga aaagatggac
     2701 ggcaccgagg aactgctcgt gaagctgaac agagaggacc tgctgcggaa gcagcggacc
     2761 ttcgacaacg gcatcatccc ccaccagatc cacctgggag agctgcacgc cattctgcgg
     2821 cggcagggcg atttttaccc attcctgaag gacaaccggg aaaagatcga gaagatcctg
     2881 accttccgca tcccctacta cgtgggccct ctggccaggg gaaacagcag attcgcctgg
     2941 atgaccagaa agagcgagga aaccatcacc ccctggaact tcgaggaagt ggtggacaag
     3001 ggcgcttccg cccagagctt catcgagcgg atgaccaact tcgataagaa cctgcccaac
     3061 gagaaggtgc tgcccaagca cagcctgctg tacgagtact tcaccgtgta taacgagctg
     3121 accaaagtga aatacgtgac cgagggaatg agaaagcccg ccttcctgag cggcgagcag
     3181 aaaaaggcca tcgtggacct gctgttcaag accaaccgga aagtgaccgt gaagcagctg
     3241 aaagaggact acttcaagaa aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa
     3301 gatcggttca acgcctccct gggcacatac cacgatctgc tgaaaattat caaggacaag
     3361 gacttcctgg acaatgagga aaacgaggac attctggaag atatcgtgct gaccctgaca
     3421 ctgtttgagg acagagagat gatcgaggaa cggctgaaaa cctatgccca cctgttcgac
     3481 gacaaagtga tgaagcagct gaagcggctg agatacaccg gctggggcag gctgagccgg
     3541 aagctgatca acggcatccg ggacaagcag tccggcaaga caatcctgga tttcctgaag
     3601 tccgacggct tcgccaacag aaacttcatg cagctgatcc acgacgacag cctgaccttt
     3661 aaagaggaca tccagaaagc ccaggtgtcc ggccagggcg atagcctgca cgagcacatt
     3721 gccaatctgg ccggcagccc cgccattaag aagggcatcc tgcagacagt gaaggtggtg
     3781 gacgagctcg tgaaagtgat gggcggccac aagcccgaga acatcgtgat cgaaatggcc
     3841 agagagaacc agaccaccca gaagggacag aagaacagcc gcgagagaat gaagcggatc
     3901 gaagagggca tcaaagagct gggcagccag atcctgaaag aacaccccgt ggaaaacacc
     3961 cagctgcaga acgagaagct gtacctgtac tacctgcaga atgggcggga tatgtacgtg
     4021 gaccaggaac tggacatcaa ccggctgtcc gactacgatg tggaccatat cgtgcctcag
     4081 agctttctga aggacgactc catcgacaac aaggtgctga ccagaagcga caagaaccgg
     4141 ggcaagagcg acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg
     4201 cagctgctga acgccaagct gattacccag agaaagttcg acaatctgac caaggccgag
     4261 agaggcggcc tgagcgaact ggataaggcc ggcttcatca agagacagct ggtggaaacc
     4321 cggcagatca caaagcacgt ggcacagatc ctggactccc ggatgaacac taagtacgac
     4381 gagaatgaca agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc
     4441 gatttccgga aggatttcca gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc
     4501 cacgacgcct acctgaacgc cgtcgtggga accgccctga tcaaaaagta ccctaagctg
     4561 gaaagcgagt tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat gatcgccaag
     4621 agcgagcagg aaatcggcaa ggctaccgcc aagtacttct tctacagcaa catcatgaac
     4681 tttttcaaga ccgagattac cctggccaac ggcgagatcc ggaagcggcc tctgatcgag
     4741 acaaacggcg aaaccgggga gatcgtgtgg gataagggcc gggattttgc caccgtgcgg
     4801 aaagtgctga gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc
     4861 ttcagcaaag agtctatcct gcccaagggc aacagcgata agctgatcgc cagaaagaag
     4921 gactgggacc ctaagaagta cggcggcttc aacagcccca ccgtggccta ttctgtgctg
     4981 gtggtggcca aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa agagctactg
     5041 gggatcacca tcatggaaag aagcagcttc gagaagaatc ccatcgactt tctggaagcc
     5101 aagggctaca aagaagtgaa aaaggacctg atcatcaagc tgcctaagta ctccctgttc
     5161 gagctggaaa acggccggaa gagaatgctg gcctctgccg gcgtgctgca gaagggaaac
     5221 gaactggccc tgccctccaa atatgtgaac ttcctgtacc tggccagcca ctatgagaag
     5281 ctgaagggct cccccgagga taatgagcag aaacagctgt ttgtggaaca gcacaagcac
     5341 tacctggacg agatcatcga gcagatcagc gagttctcca agagagtgat cctggccgac
     5401 gctaatctgg acaaagtgct gtccgcctac aacaagcacc gggataagcc catcagagag
     5461 caggccgaga atatcatcca cctgtttacc ctgaccaatc tgggagcccc tgccgccttc
     5521 aagtactttg acaccaccat caaccggaag cagtacaaca ccaccaaaga ggtgctggac
     5581 gccaccctga tccggcagag catcaccggc ctgtacgaga cacggatcga cctgtctcag
     5641 ctgggaggcg actctggcgg ctcaaaaaga accgccgacg gcagcgaatt cgagcccaag
     5701 aagaagagga aagtcgatta caaagacgat gacgataagg gatccggcgc aacaaacttc
     5761 tctctgctga aacaagccgg agatgtcgaa gagaatcctg gaccgatggc caagcctttg
     5821 tctcaagaag aatccaccct cattgaaaga gcaacggcta caatcaacag catccccatc
     5881 tctgaagact acagcgtcgc cagcgcagct ctctctagcg acggccgcat cttcactggt
     5941 gtcaatgtat atcattttac tgggggacct tgtgcagaac tcgtggtgct gggcactgct
     6001 gctgctgcgg cagctggcaa cctgacttgt atcgtcgcga tcggaaatga gaacaggggc
     6061 atcttgagcc cctgcggacg gtgccgacag gtgcttctcg atctgcatcc tgggatcaaa
     6121 gccatagtga aggacagtga tggacagccg acggcagttg ggattcgtga attgctgccc
     6181 tctggttatg tgtgggaggg ctaa
//
