LOCUS       sequence-441073-        9541 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Adenine base editor with modified TadA variant for germline A-to-G 
            base editing in zebrafish.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9541)
  AUTHORS   Qin W, Liang F, Lin SJ, Petree C, Huang K, Zhang Y, Li L, Varshney 
            P, Mourrain P, Liu Y, Varshney GK
  TITLE     ABE-ultramax for high-efficiency biallelic adenine base editing in 
            zebrafish.
  JOURNAL   Nat Commun. 2024 Jul 4;15(1):5613. doi: 10.1038/s41467-024-49943-1.
  PUBMED    38965236
REFERENCE   2  (bases 1 to 9541)
  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: "2024-07-4"; volume: "15(1):5613. doi"; pages: " 
            10.1038/s41467-024-49943-1"
COMMENT     Sequence Label: pcs2+spCas9 ABE-Umax-nanos
FEATURES             Location/Qualifiers
     source          1..9541
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(2588..2608)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        complement(2716..2734)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2717..2734)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     promoter        complement(2769..3753)
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     rep_origin      3824..4279
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3911..3930)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4121..4142
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4305..4409
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4410..4478,4479..5270)
                     /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     complement(4628..4647)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5441..6029
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5930..5949
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6183..6200
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6317..6338
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6353..6358,6359..6376,6377..6383)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6391..6407
                     /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)."
     primer_bind     6396..6418
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6415..6431
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6415..6431
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6450..6470
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6452..6470
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     polyA_signal    6583..6717
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6620..6639)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6674..6693
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(7392..7412)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     RBS             7507..7515
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
ORIGIN
        1 tcagtctacc ccagccagtg tagcgccttc tcttcagctg tttcatcacc ttgtcatcga
       61 acagatgagc ataggttttc agtctttcct cgatcattcc tctgtcctca aacagggtca
      121 gtgtcagcac aatgtcttcc aggatatcct cattttcctc gttgtccagg aagtctttat
      181 ccttaatgat cttcagcaga tcgtggtaag cgcccaggct tgcgttaaat ctgtcttcga
      241 ctccgctgat ctccacggaa tcgaagcatt caattttctt gaagtagtcc tccttcagct
      301 gcttgacagt cactttcctg ttggttttaa acagcagatc cacgattgct ttcttctgct
      361 ctcctgacag gaaggctggc tttctcatgc cctcggtgac gtatttcacc tttgtcagtt
      421 cgttatagac agtgaaatac tcgtacagca gggagtgctt aggcagcact ttctcattgg
      481 gcaggttttt gtcaaaatta gtcattctct cgatgaagga ctgtgcagag gcgcccttat
      541 cgaccacttc ctcgaagttc caaggagtaa tggtttcctc ggatttgcga gtcatccagg
      601 caaatctgct gtttccgcga gccagtggac ccacatagta agggattctg aaggtcagaa
      661 ttttttcgat cttctcgcgg ttatctttca gaaaggggta gaagtcttcc tggcgcctca
      721 ggatagcgtg cagctctccc agatgaatct ggtgtgggat ggaaccgtta tcaaatgtgc
      781 gctgcttcct cagcaggtcc tctctattca gtttcaccag cagttcctca gttccgtcca
      841 tcttctccag aataggtttg ataaacttgt agaattcctc ctgtgatgcg cctccgtcga
      901 tataaccggc gtagccgttc ttggactgat caaagaaaat ttccttgtac ttctctggca
      961 gctgctgcct gaccagagcc ttcagcaggg tcagatcctg atggtgctcg tcatatctct
     1021 tgatcatgct ggcactcaga ggagcttttg taatctcgct gttcactctc agaatatctg
     1081 acagcaggat tgcgtcggac agatttttgg cagccaggaa caggtcagcg tactgatctc
     1141 cgatctgtgc cagcaggtta tccaggtcat cgtcatatgt gtcctttgac agctgcagtt
     1201 tggcgtcctc agccagatca aaattgcttt tgaagttagg agtcaggccc agactcagag
     1261 cgatcagatt tccaaacaga ccgttcctct tttctccggg cagctgtgca atcaggttct
     1321 ccagtctgcg tgacttggac agcctggctg acaggatagc ttttgcgtcc actctggaag
     1381 cattaatagg gttttcctca aacagctgat tgtaaatctg gaccagctgg atgaacagct
     1441 tgtccacatc ggaattgtcg gggttcagat cgccctcgat cagaaaatga cccctgaact
     1501 taatcatgtg ggccagagcc agatagatca gtctcaggtc ggctttatca gtagagtcag
     1561 ccagtttctt gcgcagatga tagatggttg ggtatttttc gtgataggcg acctcgtcca
     1621 caatgtttcc aaagatagga tggcgctcgt gtttcttatc ttcctcgacc aggaaacttt
     1681 cctccagcct atgaaagaat gaatcgtcca ccttagccat ttcgttagag aaaatctcct
     1741 gcaggtaaca gattctattc ttccttctgg tatagcgcct tcttgctgtt cttttcaggc
     1801 gtgttgcttc ggcagtctct ccagaaccga acagcagggc tccaatcaga tttttcttga
     1861 tgctgtgcct gtcggtgtta cccagcacct taaatttctt gctagggacc ttgtagtcgt
     1921 ctgtaatcac tgcccaaccc acggagttag ttccaatagc caggccgatg ctatacttct
     1981 tatcactgcc gccagaggag cctccactgg actctggagt ggcagactct gatgtcccag
     2041 gtgtttcgct tccactggag cctccgctgg aacctccgct attaatggaa gactgggctt
     2101 tcttctgagc gttgaaaacc cgtcttggca ttctgtagaa atcgcacagc agagcggcac
     2161 attcgtcggc cagaatgcct tcggtgatct cgaccctatg attcattcct gggtagttca
     2221 gaacattcat cagactgccg gcagctccgc gctttgagtt tctaactcca aacacgactc
     2281 taccgatcct actgtggatc atggctcctg cgcacatcac acaaggctcg aaggtgctat
     2341 acagtgtggc gtcaatgagt ctatagttct gcatgaccag tcctccttgc ctgagtgcca
     2401 taatctctgc gtgtgcggtt gggtcatgca gtccaattgc gcggttccat ccctcgccta
     2461 taaccctgtt attcaggacc agcactgcgc ccacaggaac ctccctctca tctctagccc
     2521 tctttgcgag tgtgagagca tgtctcatcc agtattcgtg tgaaaactcg acttcgctca
     2581 tgcttcccac ctttctcttt ttcttggggc tctcgaactc ggatccatct gcagttcttt
     2641 tcatggtggc gtcgagttag atctggtggc gtcgacattg gatcggatcc tgcaaaaaga
     2701 acaagtagct tgtattctat agtgtcacct aaatcaagct tgctccgacg tccccaggca
     2761 gaatggcggt tccctaaacg agcattgctt atatagacct cccattaggc acgcctaccg
     2821 cccatttacg tcaatggaac gcccatttgc gtcattgccc ctccccattg acgtcaatgg
     2881 ggatgtactt ggcagccatc gcgggccatt taccgccatt gacgtcaatg ggagtactgc
     2941 caatgtaccc tggcgtactt ccaatagtaa tgtacttgcc aagttactat taatagatat
     3001 tgatgtactg ccaagtgggc catttaccgt cattgacgtc aatagggggc gtgagaacgg
     3061 atatgaatgg gcaatgagcc atcccattga cgtcaatggt gggtggtcct attgacgtca
     3121 atgggcattg agccaggcgg gccatttacc gtaattgacg tcaatggggg aggcgccata
     3181 tacgtcaata ggaccgccca tatgacgtca ataggaaaga ccatatatag agaccattga
     3241 cgtcaatggg ggagtggcta tgggcggtat taggaagccc catatatggt atatgggacc
     3301 gcccattggg aggggctatc tacgtcaata ggaaaaccca tatatggaat actatatggc
     3361 atagggccaa tacatagtat tgaacctggc caatagccat attggcatag ggccatattg
     3421 gatattgcct atatattgat cctggcatat agccaatatg gccgccatta ttggcaccat
     3481 gccaattcaa tatggcggac ttggcacagg gccaagaccc ctcctcaaac ttggcaccgt
     3541 gccaagtaga cccctcccca gttggcacag tgccaggtcc gccatattga attggcatgg
     3601 tgccaagtcc gccatattga attggctggg gtccagatcc accatattga attggcacga
     3661 gtccatatac gccatattga attggcacag gtccagatcc accatattga attggcatga
     3721 gtccatatac gccatattga attggcggtc gactggcgta atagcgaaga ggcccgcacc
     3781 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc
     3841 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     3901 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     3961 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     4021 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     4081 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     4141 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     4201 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacgaa
     4261 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     4321 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     4381 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     4441 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     4501 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     4561 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     4621 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     4681 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     4741 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     4801 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     4861 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     4921 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     4981 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     5041 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     5101 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     5161 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     5221 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     5281 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     5341 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     5401 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     5461 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     5521 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     5581 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     5641 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     5701 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     5761 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     5821 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     5881 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     5941 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     6001 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     6061 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     6121 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     6181 gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg
     6241 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     6301 gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact
     6361 ttatgctccc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     6421 acagctatga ccatgattac gccaagcgcg caattaaccc tcactaaagg gaacaaaagc
     6481 tgggtaccgg gcccaatgca ttggcgccgc ggccgcgaat taaaaaacct cccacacctc
     6541 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct
     6601 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     6661 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatctac
     6721 gtaatacgac tcactatagt tctagatgtt tatattttcc tcacattttt cactccatca
     6781 caagtcaaaa atcacacact gacataaaat gatgaaagta gtacatgaat tttagcaagc
     6841 gacatgaaaa tatggcgatt atgtttttca agctgaattt aaatgcaatg gagaaaactt
     6901 tttttaaagg ttacttatac aaaacaagcc gttttcaata ataaaatcac taaaatctaa
     6961 atattaacat tgaatttttt gtttaaatac aacaccataa aatctgacat ttaaaaggag
     7021 gactaaacac acattgtatt taccaaataa gcatgaaagg gtttgttttt gagtgcggtt
     7081 gcgcctgtag cgagcctgct ggcttgtgta cgagtttgtt accaagtgaa aacacaacac
     7141 cagtgcacac aaaataaaac acagcaaaca cacacatata cacacacaca aaaaaagtgc
     7201 aaaatcaaac agtgaacgca cacatgcatg caaactcgca atccgccgcg tatcaaccaa
     7261 acaaacaatt aaagagttaa aacttaaaca ttaaaacctg cggtaaaaag tgtttcttca
     7321 aatctcccgg agcatcaatg tccgctaccg gtgagtaata cgactcacta tagttctaga
     7381 ttccgcggtc acacctttct cttcttctta ggagatctca ccggtgagtc gccacccagc
     7441 tgagacaggt caatccttgt ttcgtacagg ccggtaatac tctgatggat cagtgtagcg
     7501 tccagcacct ccttagtgga ggtgtacctc tttctatcga ttgtagtgtc gaagtacttg
     7561 aaagctgcag gtgcgcccag gtttgtcaga gtgaacagat gaatgatatt ttcggcctgc
     7621 tccctgatgg gcttatctct gtgtttgttg taagcagaca ggaccttatc cagatttgcg
     7681 tcggccagga tcactctttt agagaactcg ctaatctgtt caatgatctc gtccagataa
     7741 tgcttgtgct gctccacaaa cagctgtttc tgttcgttat cctctggaga tcctttcagc
     7801 ttctcgtagt ggcttgccag atacaggaag ttcacgtact tagaaggcag ggccagttca
     7861 ttgcctttct gcagctctcc agcgcttgcc agcatgcgtt tcctgccgtt ttccagctca
     7921 aacagactgt acttgggcag tttaatgatc aggtctttct tcacttcctt ataacctttg
     7981 gcctccagaa aatcgattgg gttcttctcg aaagagcttc tctccataat tgtgataccc
     8041 agcagctcct tgacagattt cagtttcttg cttttgccct tctccacctt agcgaccacc
     8101 aggacggagt atgccacagt gggagagtca aagcctccat acttcttagg gtcccagtct
     8161 ttctttctag cgatcagctt atcagagttg cgttttggca gaatactttc ttttgagaaa
     8221 ccgcctgtct gcacctcagt tttcttgacg atattcactt gaggcatgct caggaccttc
     8281 ctcacggttg caaaatctct gcctttgtcc cacacgattt cgcctgtctc tccgtttgtc
     8341 tcaatcaggg gccttttgcg gatttctcca ttggccaggg tgatctctgt cttaaagaaa
     8401 ttcatgatgt ttgagtaaaa gaaatacttg gcagtagctt ttccaatttc ctgctcggac
     8461 ttggcgatca tttttctgac atcgtacacc ttatagtctc cgtacacgaa ttcgctctcc
     8521 agttttgggt atttcttgat cagtgcggta ccgaccacgg cgttcaggta tgcgtcatgg
     8581 gcatggtgat aattgttaat ctcgcggacc ttataaaact ggaaatcttt cctaaagtcg
     8641 ctgaccagtt tactcttcag ggtaatgacc ttcacttctc tgatcagttt gtcattctca
     8701 tcgtactttg tgttcatgcg actatccaga atctgtgcga cgtgtttagt gatctggcgt
     8761 gtttccacca gctgcctctt gatgaatcca gctttgtcca gttctgacag tccaccgcgc
     8821 tctgcttttg tcaggttatc aaacttcctc tgagtgatca gtttggcatt cagcagctgt
     8881 ctccagtagt tcttcatttt cttgaccact tcctcactag ggacattatc tgattttcct
     8941 ctgtttttgt cggacctggt cagcaccttg ttgtcaatgc tgtcatcttt gatgaaactc
     9001 tgggggacga tatgatccac gtcgtaatca gacagcctgt taatatccag ctcctggtcc
     9061 acgtacatat ctcttccatt ctgcagatag tacagataca gcttctcatt ctgcagctgt
     9121 gtgttttcca ctgggtgctc tttcaggatc tggctaccca gctccttaat gccttcctcg
     9181 attcttttca tcctctctct actgttcttc tgacccttct gggttgtctg attttcgcga
     9241 gccatctcaa tcacgatgtt ttcgggctta tggcccatga ctttcaccag ctcatcgacg
     9301 attttcactg tctgcagaat gcctttcttg atggctgggc ttccagccag gtttgcaatc
     9361 tgctcatgca gggagtggcc ctgtccagac acctgagcct tctggatgtc ctctttgaag
     9421 gtcagactgt catcgtgaat cagctgcata aaattgcggt tggcgaagcc atcacttttc
     9481 agaaagtcca ggattgtctt tcctgactgt ttatccctaa tgccgttgat cagcttcctg
     9541 c
//
