LOCUS       sequence-394828-        7626 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of N-terminally fused Nme2-evoFERNY-nt.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7626)
  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 7626)
  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: pEJS2191_Nme2-evoFERNY-nt
FEATURES             Location/Qualifiers
     source          1..7626
                     /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"
     CDS             4554..4802
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             4845..4865
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4874..4891
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4914..4931)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4920..5144
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5215..5231)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5215..5231)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5228..5250)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5239..5255
                     /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(5263..5269,5270..5287,5288..5293))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5308..5329
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5446..5463)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5617..6205)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5697..5716)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6376..7167,7168..7236))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFSAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6999..7018
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7237..7341)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7420..7439)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(7611..7626,1..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 ggaaagtcag ttttgagagg aactacgacc
      721 cccgggagct gagaaaggag acatacctgc tgtatgagat caagtggggc aagtccggca
      781 agctgtggag gcactggtgc cagaacaatc gcacacagca cgccgaggtg tacttcctgg
      841 agaacatctt taatgcccgg agattcaatc catctaccca ctgtagcatc acatggtatc
      901 tgagctggtc cccctgcgcc gagtgttctc agaagatcgt ggatttcctg aaggagcacc
      961 ctaacgtgaa tctggagatc tatgtggccc ggctgtacta tccagagaac gagaggaata
     1021 ggcagggcct gcgggatctg gtgaattccg gcgtgaccat cagaatcatg gacctgccag
     1081 attacaacta ttgctggaag accttcgtga gcgatcaggg aggcgacgag gattactggc
     1141 caggacactt cgccccttgg atcaagcagt atagcctgaa gctgtctggc ggatctagcg
     1201 gaggatcctc tggcagcgag acaccaggaa caagcgagtc agcaacacca gagagcagtg
     1261 gcggcagcag cggcggcagc gccgccttca agcctaaccc aatcaattac atcctgggac
     1321 tggccatcgg aatcgcatcc gtgggatggg ctatggtgga gatcgacgag gaggagaatc
     1381 ctatccggct gatcgatctg ggcgtgagag tgtttgagag ggccgaggtg ccaaagaccg
     1441 gcgattctct ggctatggcc cggagactgg cacggagcgt gaggcgcctg acacggagaa
     1501 gggcacacag gctgctgagg gcacgccggc tgctgaagag agagggcgtg ctgcaggcag
     1561 cagacttcga tgagaatggc ctgatcaaga gcctgccaaa caccccctgg cagctgagag
     1621 cagccgccct ggacaggaag ctgacaccac tggagtggtc tgccgtgctg ctgcacctga
     1681 tcaagcaccg cggctacctg agccagcgga agaacgaggg agagacagca gacaaggagc
     1741 tgggcgccct gctgaaggga gtggccaaca atgcccacgc cctgcagacc ggcgatttca
     1801 ggacacctgc cgagctggcc ctgaataagt ttgagaagga gtccggccac atcagaaacc
     1861 agaggggcga ctatagccac accttctccc gcaaggatct gcaggccgag ctgatcctgc
     1921 tgttcgagaa gcagaaggag tttggcaatc cacacgtgag cggaggcctg aaggagggaa
     1981 tcgagaccct gctgatgaca cagaggcctg ccctgtccgg cgacgcagtg cagaagatgc
     2041 tgggacactg caccttcgag cctgcagagc caaaggccgc caagaacacc tacacagccg
     2101 agcggtttat ctggctgaca aagctgaaca atctgagaat cctggagcag ggatccgaga
     2161 ggccactgac cgacacagag agggccaccc tgatggatga gccttaccgg aagtctaagc
     2221 tgacatatgc ccaggccaga aagctgctgg gcctggagga caccgccttc tttaagggcc
     2281 tgagatacgg caaggataat gccgaggcct ccacactgat ggagatgaag gcctatcacg
     2341 ccatctctcg cgccctggag aaggagggcc tgaaggacaa gaagtccccc ctgaacctga
     2401 gctccgagct gcaggatgag atcggcaccg ccttctctct gtttaagacc gacgaggata
     2461 tcacaggccg cctgaaggac agggtgcagc ctgagatcct ggaggccctg ctgaagcaca
     2521 tctctttcga taagtttgtg cagatcagcc tgaaggccct gagaaggatc gtgccactga
     2581 tggagcaggg caagcggtac gacgaggcct gcgccgagat ctacggcgat cactatggca
     2641 agaagaacac agaggagaag atctatctgc cccctatccc tgccgacgag atcagaaatc
     2701 ctgtggtgct gagggccctg tcccaggcaa gaaaagtgat caacggagtg gtgcgccggt
     2761 acggatctcc agcccggatc cacatcgaga ccgccagaga agtgggcaag agcttcaagg
     2821 accggaagga gatcgagaag agacaggagg agaatcgcaa ggatcgggag aaggccgccg
     2881 ccaagtttag ggagtacttc cctaactttg tgggcgagcc aaagtctaag gacatcctga
     2941 agctgcgcct gtacgagcag cagcacggca agtgtctgta tagcggcaag gagatcaatc
     3001 tggtgcggct gaacgagaag ggctatgtgg agatcgatca cgccctgcct ttctccagaa
     3061 cctgggacga ttcttttaac aataaggtgc tggtgctggg cagcgagaac cagaataagg
     3121 gcaatcagac accatacgag tatttcaatg gcaaggacaa ctccagggag tggcaggagt
     3181 tcaaggcccg cgtggagacc tctagatttc ccaggagcaa gaagcagcgg atcctgctgc
     3241 agaagttcga cgaggatggc tttaaggagt gcaacctgaa tgacaccaga tacgtgaacc
     3301 ggttcctgtg ccagtttgtg gccgatcaca tcctgctgac cggcaagggc aagagaaggg
     3361 tgttcgcctc taatggccag atcacaaacc tgctgagggg attttgggga ctgaggaagg
     3421 tgcgggcaga gaatgacaga caccacgcac tggatgcagt ggtggtggca tgcagcaccg
     3481 tggcaatgca gcagaagatc acaagattcg tgaggtataa ggagatgaac gcctttgacg
     3541 gcaagaccat cgataaggag acaggcaagg tgctgcacca gaagacccac ttcccccagc
     3601 cttgggagtt ctttgcccag gaagtgatga tccgggtgtt cggcaagcca gacggcaagc
     3661 ctgagtttga ggaggccgat accccagaga agctgaggac actgctggca gagaagctgt
     3721 ctagcaggcc agaggcagtg cacgagtacg tgaccccact gttcgtgtcc agggcaccca
     3781 atcggaagat gtctggcgcc cacaaggaca cactgagaag cgccaagagg tttgtgaagc
     3841 acaacgagaa gatctccgtg aagagagtgt ggctgaccga gatcaagctg gccgatctgg
     3901 agaacatggt gaattacaag aacggcaggg agatcgagct gtatgaggcc ctgaaggcaa
     3961 ggctggaggc ctacggagga aatgccaagc aggccttcga cccaaaggat aacccctttt
     4021 ataagaaggg aggacagctg gtgaaggccg tgcgggtgga gaagacccag gagagcggcg
     4081 tgctgctgaa taagaagaac gcctacacaa tcgccgacaa tggcgatatg gtgagagtgg
     4141 acgtgttctg taaggtggat aagaagggca agaatcagta ctttatcgtg cctatctatg
     4201 cctggcaggt ggccgagaac atcctgccag acatcgattg caagggctac agaatcgacg
     4261 atagctatac attctgtttt tccctgcaca agtatgacct gatcgccttc cagaaggatg
     4321 agaagtccaa ggtggagttt gcctactata tcaattgcga ctcctctaac ggcaggttct
     4381 acctggcctg gcacgataag ggcagcaagg agcagcagtt tcgcatctcc acccagaatc
     4441 tggtgctgat ccagaagtat caggtgaacg agctgggcaa ggagatcagg ccatgtcggc
     4501 tgaagaagcg cccacccgtg cggagcggag gatccggagg atctggaggc agcaccaacc
     4561 tgtctgacat catcgagaag gagacaggca agcagctggt catccaggag agcatcctga
     4621 tgctgcccga agaagtcgaa gaagtgatcg gaaacaagcc tgagagcgat atcctggtcc
     4681 ataccgccta cgacgagagt accgacgaaa atgtgatgct gctgacatcc gacgccccag
     4741 agtataagcc ctgggctctg gtcatccagg attccaacgg agagaacaaa atcaaaatgc
     4801 tgtctggcgg ctcaaaaaga accgccgacg gcagcgaatt cgagcccaag aagaagagga
     4861 aagtctaacc ggtcatcatc accatcacca ttgagtttaa acccgctgat cagcctcgac
     4921 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     4981 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     5041 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     5101 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     5161 aaccagctgg ggctcgatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     5221 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     5281 cataaagtgt aaagcctagg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     5341 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5401 acgcgcgggg agaggcggtt tgcgtattgg gcgcacttcc gcttcctcgc tcactgactc
     5461 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5521 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5581 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5641 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5701 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5761 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     5821 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     5881 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     5941 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     6001 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     6061 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     6121 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6181 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacac
     6241 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6301 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6361 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6421 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6481 cttaccatct ggccccagtg ctgcaatgat accgcgggac ccacgctcac cggctccaga
     6541 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6601 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6661 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6721 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     6781 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     6841 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     6901 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     6961 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     7021 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     7081 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7141 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcagaaaa
     7201 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7261 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7321 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
     7381 gggagatcga tctcccgatc ccctagggtc gactctcagt acaatctgct ctgatgccgc
     7441 atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag
     7501 caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag
     7561 ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt gacattgatt
     7621 attgac
//
