LOCUS       sequence-404875-        7540 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector for cytosine base editing using miniSdd6 at HPD-T2 target
            site in mouse.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7540)
  AUTHORS   Huang J, Lin Q, Fei H, He Z, Xu H, Li Y, Qu K, Han P, Gao Q, Li B, 
            Liu G, Zhang L, Hu J, Zhang R, Zuo E, Luo Y, Ran Y, Qiu JL, Zhao KT,
            Gao C
  TITLE     Discovery of deaminase functions by structure-based protein 
            clustering.
  JOURNAL   Cell. 2023 Jul 20;186(15):3182-3195.e14. doi: 
            10.1016/j.cell.2023.05.041. Epub 2023 Jun 27.
  PUBMED    37379837
REFERENCE   2  (bases 1 to 7540)
  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: 
            "10.1016/j.cell.2023.05.041"; journalName: "Cell"; date: 
            "2023-07-20- 20"; volume: "186"; issue: "15"; pages: "3182-3195.e14"
COMMENT     Sequence Label: pAAV-EFS-SaKKH-spA-miniU6-miniSdd6-MmHPD-T2
FEATURES             Location/Qualifiers
     source          1..7540
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        186..397
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      422..431
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             467..487
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4199..4447
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             4490..4510
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_RNA        complement(4585..4660)
                     /label=Sa gRNA scaffold
                     /note="guide RNA scaffold for the Staphylococcus aureus 
                     CRISPR/Cas9 system"
     primer_bind     complement(4773..4793)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     repeat_region   4802..4942
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4802..4931
                     /label=AAV2 ITR
     rep_origin      5017..5472
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5104..5123)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5314..5335
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5489..5508)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5608..5630
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5668..5686)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5754..5858
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5859..5927,5928..6719)
                     /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(6077..6096)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6890..7479
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7380..7399
                     /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 gataggtctt gaaaggagtg ggaattggct ccggtgcccg
      181 tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa
      241 ttgatccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg
      301 gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa
      361 cgttcttttt cgcaacgggt ttgccgccag aacacaggac cggttctaga gcgctaagct
      421 tgccaccatg gccaaacgga cagccgacgg aagcgagttc gagtcaccaa agaagaagcg
      481 gaaagtccct gcctccaagc caagtcgctt caaacccatg agaacagacc cagccaaggc
      541 agatgaaatc cgccctcacg tgggcaagga cagagctgtg gccaccctct gggatgcaga
      601 cggaaacaga gttctgggcc tgcacagcgc cgacgatgac ggccctgctg ccaccgccgc
      661 ctggaagcct ccctggcgtg actatgtgag gctgcgccgg cacgtggaag cccacgccgc
      721 cgcccggatg caccaggacg ggcacaaaac catggtaatg tatatcaacc tcccaccctg
      781 taaatacttt gatggctgca agttgaacct ggaggacata ctgcctaagg gcagcaccct
      841 gtggatgcac agggtattcc aaaatggagg gaccaagata taccagttca atggtacagg
      901 aagagcttac gtctccggag gatctagcgg aggctcctct ggctctgaga cacctggcac
      961 aagcgagagc gcaacacctg aaagcagcgg gggcagcagc ggggggtcag ggaagcgaaa
     1021 ttacattctg gggctggcca ttggcattac atcagtgggc tatggcatca ttgactacga
     1081 gacaagggac gtgatcgacg ccggcgtgag actgttcaag gaggccaacg tggagaacaa
     1141 tgagggccgg agatccaaga ggggagcaag gcgcctgaag cggagaaggc gccacagaat
     1201 ccagagagtg aagaagctgc tgttcgatta caacctgctg accgaccact ccgagctgtc
     1261 tggcatcaat ccttatgagg ccagagtgaa gggcctgtcc cagaagctgt ctgaggagga
     1321 gtttagcgcc gccctgctgc acctggcaaa gaggagaggc gtgcacaacg tgaatgaggt
     1381 ggaggaggac accggcaacg agctgtccac aaaggagcag atcagccgca attccaaggc
     1441 cctggaggag aagtatgtgg ccgagctgca gctggagcgg ctgaagaagg atggcgaggt
     1501 gaggggctcc atcaatcgct tcaagacctc tgactacgtg aaggaggcca agcagctgct
     1561 gaaggtgcag aaggcctacc accagctgga tcagtccttt atcgatacat atatcgacct
     1621 gctggagaca aggcgcacat actatgaggg accaggagag ggctctccct tcggctggaa
     1681 ggacatcaag gagtggtacg agatgctgat gggccactgc acctattttc cagaggagct
     1741 gagaagcgtg aagtacgcct ataacgccga tctgtacaac gccctgaatg acctgaacaa
     1801 cctggtcatc accagggatg agaacgagaa gctggagtac tatgagaagt tccagatcat
     1861 cgagaacgtg ttcaagcaga agaagaagcc tacactgaag cagatcgcca aggagatcct
     1921 ggtgaacgag gaggacatca agggctaccg cgtgacctcc acaggcaagc cagagttcac
     1981 caatctgaag gtgtatcacg atatcaagga catcacagcc cggaaggaga tcatcgagaa
     2041 cgccgagctg ctggatcaga tcgccaagat cctgaccatc tatcagagct ccgaggacat
     2101 ccaggaggag ctgaccaacc tgaatagcga gctgacacag gaggagatcg agcagatcag
     2161 caatctgaag ggctacaccg gcacacacaa cctgagcctg aaggccatca atctgatcct
     2221 ggatgagctg tggcacacaa acgacaatca gatcgccatc tttaaccggc tgaagctggt
     2281 gccaaagaag gtggacctgt cccagcagaa ggagatccca accacactgg tggacgattt
     2341 catcctgtct cccgtggtga agcggagctt catccagagc atcaaagtga tcaacgccat
     2401 catcaagaag tacggcctgc ccaatgatat catcatcgag ctggccaggg agaagaactc
     2461 caaggacgcc cagaagatga tcaatgagat gcagaagagg aaccgccaga ccaatgagcg
     2521 gatcgaggag atcatcagaa ccacaggcaa ggagaacgcc aagtacctga tcgagaagat
     2581 caagctgcac gatatgcagg agggcaagtg tctgtattct ctggaggcca tccctctgga
     2641 ggacctgctg aacaatccat tcaactacga ggtggatcac atcatccccc ggagcgtgag
     2701 cttcgacaat tcttttaaca ataaggtgct ggtgaagcag gaggagaaca gcaagaaggg
     2761 caataggacc cctttccagt acctgtctag ctccgattct aagatcagct acgagacatt
     2821 caagaagcac atcctgaatc tggccaaggg caagggccgc atcagcaaga ccaagaagga
     2881 gtacctgctg gaggagcggg acatcaacag attctccgtg cagaaggact tcatcaaccg
     2941 gaatctggtg gacaccagat acgccacacg cggcctgatg aatctgctgc ggtcttattt
     3001 cagagtgaac aatctggatg tgaaggtgaa gagcatcaac ggcggcttca cctcctttct
     3061 gcggagaaag tggaagttta agaaggagcg caacaagggc tataagcacc acgccgagga
     3121 tgccctgatc atcgccaatg ccgacttcat ctttaaggag tggaagaagc tggacaaggc
     3181 caagaaagtg atggagaacc agatgttcga ggagaagcag gccgagagca tgcccgagat
     3241 cgagacagag caggagtaca aggagatttt catcacacct caccagatca agcacatcaa
     3301 ggacttcaag gactacaagt attctcacag ggtggataag aagcccaacc gcaagctgat
     3361 caatgacacc ctgtatagca cacggaagga cgataagggc aataccctga tcgtgaacaa
     3421 tctgaacggc ctgtacgaca aggataatga caagctgaag aagctgatca acaagtctcc
     3481 cgagaagctg ctgatgtacc accacgatcc tcagacatat cagaagctga agctgatcat
     3541 ggagcagtac ggcgacgaga agaacccact gtataagtac tatgaggaga caggcaacta
     3601 cctgacaaag tatagcaaga aggataatgg ccccgtgatc aagaagatca agtactatgg
     3661 caacaagctg aatgcccacc tggacatcac cgacgattac cctaactctc gcaataaggt
     3721 ggtgaagctg agcctgaagc cataccggtt cgacgtgtac ctggacaacg gcgtgtataa
     3781 gtttgtgaca gtgaagaatc tggatgtgat caagaaggag aactactatg aggtgaacag
     3841 caagtgctac gaggaggcca agaagctgaa gaagatcagc aaccaggccg agttcatcgc
     3901 ctctttttac aagaatgacc tgatcaagat caatggcgag ctgtatagag tgatcggcgt
     3961 gaacaatgat ctgctgaaca gaatcgaagt gaatatgatc gacatcacct acagggagta
     4021 tctggagaac atgaatgata agaggccccc tcacatcatc aagaccatcg cctctaagac
     4081 acagagcatc aagaagtaca gcacagacat cctggggaac ctgtatgaag tcaagagcaa
     4141 gaaacatcct cagattatca agaaaggctc tggcggctca ggaggatccg gaggcagcac
     4201 caacctgtct gacatcatcg agaaggagac aggcaagcag ctggtcatcc aggagagcat
     4261 cctgatgctg cccgaagaag tcgaagaagt gatcggaaac aagcctgaga gcgatatcct
     4321 ggtccatacc gcctacgacg agagtaccga cgaaaatgtg atgctgctga catccgacgc
     4381 cccagagtat aagccctggg ctctggtcat ccaggattcc aacggagaga acaaaatcaa
     4441 aatgctgtct ggcggctcaa aaagaaccgc cgacggcagc gaattcgagc ccaagaagaa
     4501 gaggaaagtc taagctagca ataaaagatc tttattttca ttagatctgt gtgttggttt
     4561 tttgtgtaag cggtaccaaa aaaatctcgc caacaagttg acgagataaa cacggcattt
     4621 tgccttgtta tagtagattc tgtttccaga gtactataac tggttgtgac gttgaatgac
     4681 tcggtgtttc gtcctttcca caagatatat aaagccaaga aatcgaaata ctttcaagtt
     4741 acggtaagct atcgtatatg caaatatgaa ggaatcatgg gaaataggcc ctcgcggccg
     4801 caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag
     4861 gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag
     4921 cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc
     4981 ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa
     5041 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc
     5101 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     5161 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     5221 aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     5281 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     5341 cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct
     5401 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     5461 cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     5521 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     5581 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     5641 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     5701 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     5761 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     5821 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     5881 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     5941 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     6001 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     6061 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     6121 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     6181 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     6241 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     6301 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     6361 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     6421 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     6481 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     6541 ttattgctga taaatctgga gccggtgagc gtggaagccg cggtatcatt gcagcactgg
     6601 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     6661 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     6721 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     6781 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     6841 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     6901 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaaac caccgctacc agcggtggtt
     6961 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     7021 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     7081 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     7141 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     7201 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     7261 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     7321 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     7381 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     7441 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     7501 ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
