LOCUS       sequence-394815-        7416 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of N-terminally fused Nme2.Smu-ABE8e-nt.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7416)
  AUTHORS   Bamidele N, Zhang H, Dong X, Cheng H, Gaston N, Feinzig H, Cao H, 
            Kelly K, Watts JK, Xie J, Gao G, Sontheimer EJ
  TITLE     Domain-inlaid Nme2Cas9 adenine base editors with improved activity 
            and targeting scope.
  JOURNAL   Nat Commun. 2024 Feb 17;15(1):1458. doi: 10.1038/s41467-024-45763-5.
  PUBMED    38368418
REFERENCE   2  (bases 1 to 7416)
  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: "2024-02-17"; volume: "15(1):1458. doi"; pages: " 
            10.1038/s41467-024-45763-5"
COMMENT     Sequence Label: pEJS2437_Nme2.Smu-ABE8e-nt
FEATURES             Location/Qualifiers
     source          1..7416
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     610..629
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        610..628
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             678..698
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      4173..4182
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4635..4655
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4664..4681
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4704..4721)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4710..4934
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5005..5021)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5005..5021)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5018..5040)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5029..5045
                     /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(5053..5059,5060..5077,5078..5083))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5098..5119
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5236..5253)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5407..5995)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5487..5506)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6166..6957,6958..7026))
                     /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     6789..6808
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7027..7131)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7210..7229)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(7401..7416,1..364)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagagatc
      601 cgcggccgct aatacgactc actataggga gagccgccac catgaaacgg acagccgacg
      661 gaagcgagtt cgagtcacca aagaagaagc ggaaagtcgg cggtagcggc ggaggcagcg
      721 gtggcggcag cggctctgag gtggagtttt cccacgagta ctggatgaga catgccctga
      781 ccctggccaa gagggcacgc gatgagaggg aggtgcctgt gggagccgtg ctggtgctga
      841 acaatagagt gatcggcgag ggctggaaca gagccatcgg cctgcacgac ccaacagccc
      901 atgccgaaat tatggccctg agacagggcg gcctggtcat gcagaactac agactgattg
      961 acgccaccct gtacgtgaca ttcgagcctt gcgtgatgtg cgccggcgcc atgatccact
     1021 ctaggatcgg ccgcgtggtg tttggcgtga ggaacagcaa acggggcgcc gcaggctccc
     1081 tgatgaacgt gctgaactac cccggcatga atcaccgcgt cgaaattacc gagggaatcc
     1141 tggcagatga atgtgccgcc ctgctgtgcg acttctaccg gatgcctaga caggtgttca
     1201 atgctcagaa gaaggcccag agctccatca actccggagg atctagcgga ggctcctctg
     1261 gctctgagac acctggcaca agcgagagcg caacacctga aagcagcggg ggcagcagcg
     1321 gggggtcaat ggccgccttc aagcctaacc caatcaatta catcctggga ctggccatcg
     1381 gaatcgcatc cgtgggatgg gctatggtgg agatcgacga ggaggagaat cctatccggc
     1441 tgatcgatct gggcgtgaga gtgtttgaga gggccgaggt gccaaagacc ggcgattctc
     1501 tggctatggc ccggagactg gcacggagcg tgaggcgcct gacacggaga agggcacaca
     1561 ggctgctgag ggcacgccgg ctgctgaaga gagagggcgt gctgcaggca gcagacttcg
     1621 atgagaatgg cctgatcaag agcctgccaa acaccccctg gcagctgaga gcagccgccc
     1681 tggacaggaa gctgacacca ctggagtggt ctgccgtgct gctgcacctg atcaagcacc
     1741 gcggctacct gagccagcgg aagaacgagg gagagacagc agacaaggag ctgggcgccc
     1801 tgctgaaggg agtggccaac aatgcccacg ccctgcagac cggcgatttc aggacacctg
     1861 ccgagctggc cctgaataag tttgagaagg agtccggcca catcagaaac cagaggggcg
     1921 actatagcca caccttctcc cgcaaggatc tgcaggccga gctgatcctg ctgttcgaga
     1981 agcagaagga gtttggcaat ccacacgtga gcggaggcct gaaggaggga atcgagaccc
     2041 tgctgatgac acagaggcct gccctgtccg gcgacgcagt gcagaagatg ctgggacact
     2101 gcaccttcga gcctgcagag ccaaaggccg ccaagaacac ctacacagcc gagcggttta
     2161 tctggctgac aaagctgaac aatctgagaa tcctggagca gggatccgag aggccactga
     2221 ccgacacaga gagggccacc ctgatggatg agccttaccg gaagtctaag ctgacatatg
     2281 cccaggccag aaagctgctg ggcctggagg acaccgcctt ctttaagggc ctgagatacg
     2341 gcaaggataa tgccgaggcc tccacactga tggagatgaa ggcctatcac gccatctctc
     2401 gcgccctgga gaaggagggc ctgaaggaca agaagtcccc cctgaacctg agctccgagc
     2461 tgcaggatga gatcggcacc gccttctctc tgtttaagac cgacgaggat atcacaggcc
     2521 gcctgaagga cagggtgcag cctgagatcc tggaggccct gctgaagcac atctctttcg
     2581 ataagtttgt gcagatcagc ctgaaggccc tgagaaggat cgtgccactg atggagcagg
     2641 gcaagcggta cgacgaggcc tgcgccgaga tctacggcga tcactatggc aagaagaaca
     2701 cagaggagaa gatctatctg ccccctatcc ctgccgacga gatcagaaat cctgtggtgc
     2761 tgagggccct gtcccaggca agaaaagtga tcaacggagt ggtgcgccgg tacggatctc
     2821 cagcccggat ccacatcgag accgccagag aagtgggcaa gagcttcaag gaccggaagg
     2881 agatcgagaa gagacaggag gagaatcgca aggatcggga gaaggccgcc gccaagttta
     2941 gggagtactt ccctaacttt gtgggcgagc caaagtctaa ggacatcctg aagctgcgcc
     3001 tgtacgagca gcagcacggc aagtgtctgt atagcggcaa ggagatcaat ctggtgcggc
     3061 tgaacgagaa gggctatgtg gagatcgatc acgccctgcc tttctccaga acctgggacg
     3121 attcttttaa caataaggtg ctggtgctgg gcagcgagaa ccagaataag ggcaatcaga
     3181 caccatacga gtatttcaat ggcaaggaca actccaggga gtggcaggag ttcaaggccc
     3241 gcgtggagac ctctagattt cccaggagca agaagcagcg gatcctgctg cagaagttcg
     3301 acgaggatgg ctttaaggag tgcaacctga atgacaccag atacgtgaac cggttcctgt
     3361 gccagtttgt ggccgatcac atcctgctga ccggcaaggg caagagaagg gtgttcgcct
     3421 ctaatggcca gatcacaaac ctgctgaggg gattttgggg actgaggaag gtgcgggcag
     3481 agaatgacag acaccacgca ctggatgcag tggtggtggc atgcagcacc gtggcaatgc
     3541 agcagaagat cacaagattc gtgaggtata aggagatgaa cgcctttgac ggcaagacca
     3601 tcgataagga gacaggcaag gtgctgcacc agaagaccca cttcccccag ccttgggagt
     3661 tctttgccca ggaagtgatg atccgggtgt tcggcaagcc agacggcaag cctgagtttg
     3721 aggaggccga taccccagag aagctgagga cactgctggc agagaagctg tctagcaggc
     3781 cagaggcagt gcacgagtac gtgaccccac tgttcgtgtc cagggcaccc aatcggaaga
     3841 tgtctggcgc ccacaaggac acactgagaa gcgccaagag gtttgtgaag cacaacgaga
     3901 agatctccgt gaagagagtg tggctgaccg agatcaagct ggccgatctg gagaacatgg
     3961 tgaattacaa gaacggcagg gagatcgagc tgtatgaggc cctgaaggca aggctggagg
     4021 cctacggagg aaatgccaag caggccttcg acccaaagga taaccccttt tataagaagg
     4081 gaggacagct ggtgaaggcc gtgcgggtgg agaagaccca ggagagcggc gtgctgctga
     4141 ataagaagaa cgcctacaca atcgccgaca acgccaccat ggtgcgggtg gacgtgtaca
     4201 ccaaggccgg caagaactac ctggttcctg tgtacgtgtg gcaggtggcc cagggcatct
     4261 tacccaaccg cgccgtgacc agcggcaagt ccgaggctga ctgggacctg atcgatgaga
     4321 gcttcgagtt caagttctct ctgtcccggg gagatctcgt ggaaatgatc tccaacaagg
     4381 gcagaatctt cggctactac aacggcctgg acagagccaa cggctctatt ggaattagag
     4441 agcacgacct agagaagagc aagggcaaag acggcgtgca tagagtggga gtgaaaacag
     4501 ctacagcatt taacaagtac cacgtggatc ccctgggcaa agagatccac agatgcagca
     4561 gcgaacccag acctacactg aaaatcaagt ctaagaagga ggataaaaga accgccgacg
     4621 gcagcgaatt cgagcccaag aagaagagga aagtctaacc ggtcatcatc accatcacca
     4681 ttgagtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     4741 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     4801 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     4861 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     4921 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctcgatac cgtcgacctc
     4981 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     5041 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctagg gtgcctaatg
     5101 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     5161 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     5221 gcgcacttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     5281 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     5341 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     5401 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     5461 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5521 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     5581 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     5641 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     5701 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     5761 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     5821 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     5881 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     5941 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     6001 tcctttgatc ttttctacgg ggtctgacac tcagtggaac gaaaactcac gttaagggat
     6061 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     6121 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     6181 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     6241 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     6301 accgcgggac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     6361 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     6421 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     6481 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     6541 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     6601 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     6661 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     6721 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     6781 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     6841 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     6901 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     6961 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     7021 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     7081 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     7141 ccgaaaagtg ccacctgacg tcgacggatc gggagatcga tctcccgatc ccctagggtc
     7201 gactctcagt acaatctgct ctgatgccgc atagttaaac cagtatctgc tccctgcttg
     7261 tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt
     7321 gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt
     7381 acgggccaga tatacgcgtt gacattgatt attgac
//
