LOCUS       sequence-389818-        7333 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Single AAV vector for expressing N-terminal fusion Nme2Cas9-ABE8e 
            and one miniU6 driven sgRNA targeting mouse Rosa26 gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7333)
  AUTHORS   Zhang H, Bamidele N, Liu P, Ojelabi O, Gao XD, Rodriguez T, Cheng H,
            Kelly K, Watts JK, Xie J, Gao G, Wolfe SA, Xue W, Sontheimer EJ
  TITLE     Adenine Base Editing In Vivo with a Single Adeno-Associated Virus 
            Vector.
  JOURNAL   GEN Biotechnol. 2022 Jun 1;1(3):285-299. doi: 
            10.1089/genbio.2022.0015. Epub 2022 Jun 14.
  PUBMED    35811581
REFERENCE   2  (bases 1 to 7333)
  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: 
            "10.1089/genbio.2022.0015"; journalName: "GEN Biotechnol"; date: 
            "2022-06-1- 1"; volume: "1"; issue: "3"; pages: "285-299"
COMMENT     Sequence Label: pEJS1964 All-in-one
            AAV-U1a-NmeABE-8e-2xBPSV40...
FEATURES             Location/Qualifiers
     source          1..7333
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     primer_bind     complement(369..389)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     CDS             692..712
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4625..4645
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     repeat_region   4709..4849
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4709..4838
                     /label=AAV2 ITR
     rep_origin      4924..5379
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5011..5030)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5221..5242
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5396..5415)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5548..5652
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5653..5721,5722..6513)
                     /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(5871..5890)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6684..7272
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7173..7192
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggcctcta gagtttaaac aaaaaaataa acgatgcccc ttaaagcaga
      181 agctttaagg ggcagagcgt tgcggcacat cttttcagac ggccttattg tagcaacgtt
      241 tcgggagcta caacctccca ggcccagggc ggtcctccgg tgtttcgtcc tttccacaag
      301 atatataaag ccaagaaatc gaaatacttt caagttacgg taagctatcg tatatgcaaa
      361 tatgaaggaa tcatgggaaa taggccctct ctagaatgga ggcggtacta tgtagatgag
      421 aattcaggag caaactggga aaagcaactg cttccaaata tttgtgattt ttacagtgta
      481 gttttggaaa aactcttagc ctaccaattc ttctaagtgt tttaaaatgt gggagccagt
      541 acacatgaag ttatagagtg ttttaatgag gcttaaatat ttaccgtaac tatgaaatgc
      601 tacgcatatc atgctgttca ggctccgtgg ccacgcaact catactaccg gtgccaccat
      661 gaaaagaacc gccgacggca gcgaattcga gcccaagaag aagaggaaag tcggcggtag
      721 cggcggaggc agcggtggcg gcagcggctc tgaggtggag ttttcccacg agtactggat
      781 gagacatgcc ctgaccctgg ccaagagggc acgcgatgag agggaggtgc ctgtgggagc
      841 cgtgctggtg ctgaacaata gagtgatcgg cgagggctgg aacagagcca tcggcctgca
      901 cgacccaaca gcccatgccg aaattatggc cctgagacag ggcggcctgg tcatgcagaa
      961 ctacagactg attgacgcca ccctgtacgt gacattcgag ccttgcgtga tgtgcgccgg
     1021 cgccatgatc cactctagga tcggccgcgt ggtgtttggc gtgaggaaca gcaaacgggg
     1081 cgccgcaggc tccctgatga acgtgctgaa ctaccccggc atgaatcacc gcgtcgaaat
     1141 taccgaggga atcctggcag atgaatgtgc cgccctgctg tgcgacttct accggatgcc
     1201 tagacaggtg ttcaatgctc agaagaaggc ccagagctcc atcaactccg gaggatctag
     1261 cggaggctcc tctggctctg agacacctgg cacaagcgag agcgcaacac ctgaaagcag
     1321 cgggggcagc agcggggggt caatggccgc cttcaagcct aacccaatca attacatcct
     1381 gggactggcc atcggaatcg catccgtggg atgggctatg gtggagatcg acgaggagga
     1441 gaatcctatc cggctgatcg atctgggcgt gagagtgttt gagagggccg aggtgccaaa
     1501 gaccggcgat tctctggcta tggcccggag actggcacgg agcgtgaggc gcctgacacg
     1561 gagaagggca cacaggctgc tgagggcacg ccggctgctg aagagagagg gcgtgctgca
     1621 ggcagcagac ttcgatgaga atggcctgat caagagcctg ccaaacaccc cctggcagct
     1681 gagagcagcc gccctggaca ggaagctgac accactggag tggtctgccg tgctgctgca
     1741 cctgatcaag caccgcggct acctgagcca gcggaagaac gagggagaga cagcagacaa
     1801 ggagctgggc gccctgctga agggagtggc caacaatgcc cacgccctgc agaccggcga
     1861 tttcaggaca cctgccgagc tggccctgaa taagtttgag aaggagtccg gccacatcag
     1921 aaaccagagg ggcgactata gccacacctt ctcccgcaag gatctgcagg ccgagctgat
     1981 cctgctgttc gagaagcaga aggagtttgg caatccacac gtgagcggag gcctgaagga
     2041 gggaatcgag accctgctga tgacacagag gcctgccctg tccggcgacg cagtgcagaa
     2101 gatgctggga cactgcacct tcgagcctgc agagccaaag gccgccaaga acacctacac
     2161 agccgagcgg tttatctggc tgacaaagct gaacaatctg agaatcctgg agcagggatc
     2221 cgagaggcca ctgaccgaca cagagagggc caccctgatg gatgagcctt accggaagtc
     2281 taagctgaca tatgcccagg ccagaaagct gctgggcctg gaggacaccg ccttctttaa
     2341 gggcctgaga tacggcaagg ataatgccga ggcctccaca ctgatggaga tgaaggccta
     2401 tcacgccatc tctcgcgccc tggagaagga gggcctgaag gacaagaagt cccccctgaa
     2461 cctgagctcc gagctgcagg atgagatcgg caccgccttc tctctgttta agaccgacga
     2521 ggatatcaca ggccgcctga aggacagggt gcagcctgag atcctggagg ccctgctgaa
     2581 gcacatctct ttcgataagt ttgtgcagat cagcctgaag gccctgagaa ggatcgtgcc
     2641 actgatggag cagggcaagc ggtacgacga ggcctgcgcc gagatctacg gcgatcacta
     2701 tggcaagaag aacacagagg agaagatcta tctgccccct atccctgccg acgagatcag
     2761 aaatcctgtg gtgctgaggg ccctgtccca ggcaagaaaa gtgatcaacg gagtggtgcg
     2821 ccggtacgga tctccagccc ggatccacat cgagaccgcc agagaagtgg gcaagagctt
     2881 caaggaccgg aaggagatcg agaagagaca ggaggagaat cgcaaggatc gggagaaggc
     2941 cgccgccaag tttagggagt acttccctaa ctttgtgggc gagccaaagt ctaaggacat
     3001 cctgaagctg cgcctgtacg agcagcagca cggcaagtgt ctgtatagcg gcaaggagat
     3061 caatctggtg cggctgaacg agaagggcta tgtggagatc gatcacgccc tgcctttctc
     3121 cagaacctgg gacgattctt ttaacaataa ggtgctggtg ctgggcagcg agaaccagaa
     3181 taagggcaat cagacaccat acgagtattt caatggcaag gacaactcca gggagtggca
     3241 ggagttcaag gcccgcgtgg agacctctag atttcccagg agcaagaagc agcggatcct
     3301 gctgcagaag ttcgacgagg atggctttaa ggagtgcaac ctgaatgaca ccagatacgt
     3361 gaaccggttc ctgtgccagt ttgtggccga tcacatcctg ctgaccggca agggcaagag
     3421 aagggtgttc gcctctaatg gccagatcac aaacctgctg aggggatttt ggggactgag
     3481 gaaggtgcgg gcagagaatg acagacacca cgcactggat gcagtggtgg tggcatgcag
     3541 caccgtggca atgcagcaga agatcacaag attcgtgagg tataaggaga tgaacgcctt
     3601 tgacggcaag accatcgata aggagacagg caaggtgctg caccagaaga cccacttccc
     3661 ccagccttgg gagttctttg cccaggaagt gatgatccgg gtgttcggca agccagacgg
     3721 caagcctgag tttgaggagg ccgatacccc agagaagctg aggacactgc tggcagagaa
     3781 gctgtctagc aggccagagg cagtgcacga gtacgtgacc ccactgttcg tgtccagggc
     3841 acccaatcgg aagatgtctg gcgcccacaa ggacacactg agaagcgcca agaggtttgt
     3901 gaagcacaac gagaagatct ccgtgaagag agtgtggctg accgagatca agctggccga
     3961 tctggagaac atggtgaatt acaagaacgg cagggagatc gagctgtatg aggccctgaa
     4021 ggcaaggctg gaggcctacg gaggaaatgc caagcaggcc ttcgacccaa aggataaccc
     4081 cttttataag aagggaggac agctggtgaa ggccgtgcgg gtggagaaga cccaggagag
     4141 cggcgtgctg ctgaataaga agaacgccta cacaatcgcc gacaatggcg atatggtgag
     4201 agtggacgtg ttctgtaagg tggataagaa gggcaagaat cagtacttta tcgtgcctat
     4261 ctatgcctgg caggtggccg agaacatcct gccagacatc gattgcaagg gctacagaat
     4321 cgacgatagc tatacattct gtttttccct gcacaagtat gacctgatcg ccttccagaa
     4381 ggatgagaag tccaaggtgg agtttgccta ctatatcaat tgcgactcct ctaacggcag
     4441 gttctacctg gcctggcacg ataagggcag caaggagcag cagtttcgca tctccaccca
     4501 gaatctggtg ctgatccaga agtatcaggt gaacgagctg ggcaaggaga tcaggccatg
     4561 tcggctgaag aagcgcccac ccgtgcggga ggataaaaga accgccgacg gcagcgaatt
     4621 cgagcccaag aagaagagga aagtctaaaa taaaagatct ttattttcat tagatctgtg
     4681 tgttggtttt ttgtgtaagc gcggccgcag gaacccctag tgatggagtt ggccactccc
     4741 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     4801 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg
     4861 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat
     4921 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     4981 ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg
     5041 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     5101 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg
     5161 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     5221 gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt
     5281 tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat
     5341 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa
     5401 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     5461 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgctc gagttagacg
     5521 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     5581 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     5641 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     5701 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     5761 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     5821 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     5881 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     5941 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     6001 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     6061 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     6121 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     6181 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     6241 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6301 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6361 gagcgtggaa gccgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6421 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6481 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6541 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     6601 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     6661 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     6721 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     6781 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     6841 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     6901 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     6961 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     7021 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     7081 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     7141 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     7201 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     7261 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     7321 tttgctcaca tgt
//
