LOCUS       sequence-334022-        8217 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the base editor ABE8e-NRCH in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8217)
  TITLE     ABE8e-NRCH for bacterial and human expression
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8217)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pCMV-ABE8e-NRCH
FEATURES             Location/Qualifiers
     source          1..8217
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(549..1340,1341..1409))
                     /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     1172..1191
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1410..1514)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1593..1612)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1784..2163
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2164..2367
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2318..2338
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2409..2428
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2409..2427
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2477..2497
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3092..7192
                     /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="DKKYSIGLTIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMVKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGIIPHQIHLGELHAILRRQGDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRLRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGGH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKGNSDKLIARKKDWDPKKYGGFNSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGVLQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTINRKQYNTTKEVLD
                     ATLIRQSITGLYETRIDLSQLGGD"
     CDS             7235..7255
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7264..7281
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(7304..7321)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7310..7534
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7605..7621)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7605..7621)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7618..7640)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7629..7645
                     /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(7653..7659,7660..7677,7678..7683))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7698..7719
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7836..7853)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(8007..8217,1..378))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8087..8106)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
       61 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
      121 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
      181 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
      241 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
      301 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
      361 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cactcagtgg
      421 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
      481 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
      541 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
      601 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
      661 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
      721 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
      781 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
      841 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
      901 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
      961 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     1021 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     1081 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     1141 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     1201 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     1261 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     1321 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     1381 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     1441 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     1501 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat
     1561 cgatctcccg atcccctagg gtcgactctc agtacaatct gctctgatgc cgcatagtta
     1621 agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt
     1681 taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct tagggttagg
     1741 cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gttgacattg attattgact
     1801 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
     1861 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
     1921 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
     1981 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
     2041 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
     2101 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
     2161 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
     2221 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
     2281 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
     2341 cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatccg ctagagatcc
     2401 gcggccgcta atacgactca ctatagggag agccgccacc atgaaacgga cagccgacgg
     2461 aagcgagttc gagtcaccaa agaagaagcg gaaagtctct gaggtggagt tttcccacga
     2521 gtactggatg agacatgccc tgaccctggc caagagggca cgggatgaga gggaggtgcc
     2581 tgtgggagcc gtgctggtgc tgaacaatag agtgatcggc gagggctgga acagagccat
     2641 cggcctgcac gacccaacag cccatgccga aattatggcc ctgagacagg gcggcctggt
     2701 catgcagaac tacagactga ttgacgccac cctgtacgtg acattcgagc cttgcgtgat
     2761 gtgcgccggc gccatgatcc actctaggat cggccgcgtg gtgtttggcg tgaggaactc
     2821 aaaaagaggc gccgcaggct ccctgatgaa cgtgctgaac taccccggca tgaatcaccg
     2881 cgtcgaaatt accgagggaa tcctggcaga tgaatgtgcc gccctgctgt gcgatttcta
     2941 tcggatgcct agacaggtgt tcaatgctca gaagaaggcc cagagctcca tcaactccgg
     3001 aggatctagc ggaggctcct ctggctctga gacacctggc acaagcgaga gcgcaacacc
     3061 tgaaagcagc gggggcagca gcggggggtc agacaagaag tacagcatcg gcctgaccat
     3121 cggcaccaac tctgtgggct gggccgtgat caccgacgag tacaaggtgc ccagcaagaa
     3181 attcaaggtg ctgggcaaca ccgaccggca cagcatcaag aagaacctga tcggagccct
     3241 gctgttcgac agcggcgaaa cagccgaggc cacccggctg aagagaaccg ccagaagaag
     3301 atacaccaga cggaagaacc ggatctgcta tctgcaagag atcttcagca acgagatggc
     3361 caaggtggac gacagcttct tccacagact ggaagagtcc ttcctggtgg aagaggataa
     3421 gaagcacgag cggcacccca tcttcggcaa catcgtggac gaggtggcct accacgagaa
     3481 gtaccccacc atctaccacc tgagaaagaa actggtggac agcaccgaca aggccgacct
     3541 gcggctgatc tatctggccc tggcccacat gatcaagttc cggggccact tcctgatcga
     3601 gggcgacctg aaccccgaca acagcgacgt ggacaagctg ttcatccagc tggtgcagac
     3661 ctacaaccag ctgttcgagg aaaaccccat caacgccagc ggcgtggacg ccaaggccat
     3721 cctgtctgcc agactgagca agagcagacg gctggaaaat ctgatcgccc agctgcccgg
     3781 cgagaagaag aatggcctgt tcggaaacct gattgccctg agcctgggcc tgacccccaa
     3841 cttcaagagc aacttcgacc tggccgagga tgccaaactg cagctgagca aggacaccta
     3901 cgacgacgac ctggacaacc tgctggccca gatcggcgac cagtacgccg acctgtttct
     3961 ggccgccaag aacctgtccg acgccatcct gctgagcgac atcctgagag tgaacaccga
     4021 gatcaccaag gcccccctga gcgcctctat ggtgaagaga tacgacgagc accaccagga
     4081 cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca aagagatttt
     4141 cttcgaccag agcaagaacg gctacgccgg ctacattgac ggcggagcca gccaggaaga
     4201 gttctacaag ttcatcaagc ccatcctgga aaagatggac ggcaccgagg aactgctcgt
     4261 gaagctgaac agagaggacc tgctgcggaa gcagcggacc ttcgacaacg gcattatccc
     4321 ccaccagatc cacctgggag agctgcacgc cattctgcgg cggcagggcg atttttaccc
     4381 attcctgaag gacaaccggg aaaagatcga gaagatcctg accttccgca tcccctacta
     4441 cgtgggccct ctggccaggg gaaacagcag attcgcctgg atgaccagaa agagcgagga
     4501 aaccatcacc ccctggaact tcgaggaagt ggtggacaag ggcgcttccg cccagagctt
     4561 catcgagcgg atgaccaact tcgataagaa cctgcccaac gagaaggtgc tgcccaagca
     4621 cagcctgctg tacgagtact tcaccgtgta taacgagctg accaaagtga aatacgtgac
     4681 cgagggaatg agaaagcccg ccttcctgag cggcgagcag aaaaaggcca tcgtggacct
     4741 gctgttcaag accaaccgga aagtgaccgt gaagcagctg aaagaggact acttcaagaa
     4801 aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa gatcggttca acgcctccct
     4861 gggcacatac cacgatctgc tgaaaattat caaggacaag gacttcctgg acaatgagga
     4921 aaacgaggac attctggaag atatcgtgct gaccctgaca ctgtttgagg acagagagat
     4981 gatcgaggaa cggctgaaaa cctatgccca cctgttcgac gacaaagtga tgaagcagct
     5041 gaagcggctg agatacaccg gctggggcag gctgagccgg aagctgatca acggcatccg
     5101 ggacaagcag tccggcaaga caatcctgga tttcctgaag tccgacggct tcgccaacag
     5161 aaacttcatg cagctgatcc acgacgacag cctgaccttt aaagaggaca tccagaaagc
     5221 ccaggtgtcc ggccagggcg atagcctgca cgagcacatt gccaatctgg ccggcagccc
     5281 cgccattaag aagggcatcc tgcagacagt gaaggtggtg gacgagctcg tgaaagtgat
     5341 gggcggccac aagcccgaga acatcgtgat cgaaatggcc agagagaacc agaccaccca
     5401 gaagggacag aagaacagcc gcgagagaat gaagcggatc gaagagggca tcaaagagct
     5461 gggcagccag atcctgaaag aacaccccgt ggaaaacacc cagctgcaga acgagaagct
     5521 gtacctgtac tacctgcaga atgggcggga tatgtacgtg gaccaggaac tggacatcaa
     5581 ccggctgtcc gactacgatg tggaccatat cgtgcctcag agctttctga aggacgactc
     5641 catcgacaac aaggtgctga ccagaagcga caagaaccgg ggcaagagcg acaacgtgcc
     5701 ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg cagctgctga acgccaagct
     5761 gattacccag agaaagttcg acaatctgac caaggccgag agaggcggcc tgagcgaact
     5821 ggataaggcc ggcttcatca agagacagct ggtggaaacc cggcagatca caaagcacgt
     5881 ggcacagatc ctggactccc ggatgaacac taagtacgac gagaatgaca agctgatccg
     5941 ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga aggatttcca
     6001 gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct acctgaacgc
     6061 cgtcgtggga accgccctga tcaaaaagta ccctaagctg gaaagcgagt tcgtgtacgg
     6121 cgactacaag gtgtacgacg tgcggaagat gatcgccaag agcgagcagg aaatcggcaa
     6181 ggctaccgcc aagtacttct tctacagcaa catcatgaac tttttcaaga ccgagattac
     6241 cctggccaac ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg aaaccgggga
     6301 gatcgtgtgg gataagggcc gggattttgc caccgtgcgg aaagtgctga gcatgcccca
     6361 agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag agtctatcct
     6421 gcccaagggt aacagcgata agctgatcgc cagaaagaag gactgggacc ctaagaagta
     6481 cggcggcttc aacagcccca ccgtggccta ttctgtgctg gtggtggcca aagtggaaaa
     6541 gggcaagtcc aagaaactga agagtgtgaa agagctgctg gggatcacca tcatggaaag
     6601 aagcagcttc gagaagaatc ccatcgactt tctggaagcc aagggctaca aagaagtgaa
     6661 aaaggacctg atcatcaagc tgcctaagta ctccctgttc gagctggaaa acggccggaa
     6721 gagaatgctg gcctctgccg gcgtgctgca gaagggaaac gaactggccc tgccctccaa
     6781 atatgtgaac ttcctgtacc tggccagcca ctatgagaag ctgaagggct cccccgagga
     6841 taatgagcag aaacagctgt ttgtggaaca gcacaagcac tacctggacg agatcatcga
     6901 gcagatcagc gagttctcca agagagtgat cctggccgac gctaatctgg acaaagtgct
     6961 gtccgcctac aacaagcacc gggataagcc catcagagag caggccgaga atatcatcca
     7021 cctgtttacc ctgaccaatc tgggagcccc tgccgccttc aagtactttg acaccaccat
     7081 caaccggaag caatacaaca cgaccaaaga ggtgctggac gccaccctga tccgtcagag
     7141 catcaccggc ctgtacgaga cacggatcga cctgtctcag ctgggaggtg actctggcgg
     7201 ctcaaaaaga accgccgacg gcagcgaatt cgagcccaag aagaagagga aagtctaacc
     7261 ggtcatcatc accatcacca ttgagtttaa acccgctgat cagcctcgac tgtgccttct
     7321 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     7381 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     7441 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     7501 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     7561 ggctcgatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     7621 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     7681 aaagcctagg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     7741 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     7801 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     7861 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     7921 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     7981 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     8041 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     8101 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     8161 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtagg
//
