LOCUS       sequence-437212-        6638 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV genome expressing ZFcharm Kv1 targeting Prnp; single mismatch 
            self-silencing binding site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6638)
  AUTHORS   Neumann EN, Bertozzi TM, Wu E, Serack F, Harvey JW, Brauer PP, 
            Pirtle CP, Coffey A, Howard M, Kamath N, Lenz K, Guzman K, Raymond 
            MH, Khalil AS, Deverman BE, Minikel EV, Vallabh SM, Weissman JS
  TITLE     Brainwide silencing of prion protein by AAV-mediated delivery of an 
            engineered compact epigenetic editor.
  JOURNAL   Science. 2024 Jun 28;384(6703):ado7082. doi: 
            10.1126/science.ado7082. Epub 2024 Jun 28.
  PUBMED    38935715
REFERENCE   2  (bases 1 to 6638)
  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: "Science."; 
            date: "2024-06-28"; volume: "384(6703):ado7082. doi"; pages: " 
            10.1126/science.ado7082. Epub 2024 Jun 28"
COMMENT     Sequence Label: pTMB303_ZFcharm_Kv1_Prnp_SMM
FEATURES             Location/Qualifiers
     source          1..6638
                     /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        173..384
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      392..401
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1523..1543
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2174..2194
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2243..2428
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEP"
     CDS             2438..2464
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    2486..3074
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2539..2559)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2957..2968)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3104..3152
                     /note="synthetic polyadenylation signal"
     repeat_region   3901..4041
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3901..4030
                     /label=AAV2 ITR
     rep_origin      4116..4571
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4203..4222)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4413..4434
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4588..4607)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4707..4729
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4767..4785)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4853..4957
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4958..5026,5027..5818)
                     /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(5176..5195)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5989..6577
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6478..6497
                     /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 gcggcctact agtggatcag cagaccgatt gtgggctcta gagggcagag
      181 cgcacatcgc ccacagtccc cgagaagttg gggggagggg tcggcaattg aaccggtgcc
      241 tagagaaggt ggcgcggggt aaactgggaa agtgatgtcg tgtactggct ccgccttttt
      301 cccgagggtg ggggagaacc gtatataagt gcagtagtcg ccgtgaacgt tctttttcgc
      361 aacgggtttg ccgccagaac acagcctcag cgccaccatg gccaggacca aacaaactgc
      421 cagaaagagc accgggggga aagcgccccg aaaacagctg gcgaccaaag cagcccgcaa
      481 aagcgcaccg ggaggtgcaa gcagtggggc aggaagttcc tcaggaggaa gtgcggcggg
      541 atccggaagc ggtggcgcat caagtggcgc cgcaagtagc agtgccgggt ccgcagctgg
      601 ctccgggtcc tcccctgttg agatctacaa gaccgtgtcc gcctggaagc ggcagccaat
      661 cagagtgctg agcctgttcg gcaatattga caaggaactg aagtctctgg gcttcctgga
      721 aagcggcagc cacagcgagg gcggcacact gaagtacgtg gaagatgtga ccaacgtggt
      781 gcggagagat gtggaaaagt ggggaccttt cgacctgctg tacggcagca cccagcctct
      841 gggcggaagc tgcgacaggt gtcctggctg gtacatgttc caatttcata gaatcctgca
      901 gtacgccaga cctagacagg agtctcagcc cttcttctgg atcttcatgg ataacctgct
      961 gctcacagag gacgaccaga tgaccaccgt gcggttcctg caaacagagg ccgtcaccct
     1021 gcgcgacgtg agatctagag tgctgcagaa cgccgtccgg gtgtggagca acatccccgg
     1081 tcttaaaagc aagcacgagg ctctgtcccc aaaagaggaa gagagcctgc agggccacgt
     1141 gcggaccaga tctaagctgg ccgctcctaa agtggacccc ctggtgaaga actgcctgct
     1201 gcctctgaga gagtacttta agtatttcag ccagagcagc ctgcccctgg gaggacctag
     1261 cagcggcgcc ccacctccaa gcggaggctc tccagctggc agccctacca gcacagagga
     1321 aggcacaagc gagagcgcca cacctgagag cggccctggc acctccaccg agcccagcga
     1381 gggctctgcc cctggcagcc ccgccggaag ccccacctct acagaagaag gcaccagcac
     1441 agagccttct gaaggctctg cccctggaac atccaccgag cctagcgaga agcggactgc
     1501 cgatggatct gagtttgaga gccctaagaa aaaaagaaag gttggagtgc ctgctgctat
     1561 ggccgagaga ccatttcagt gcagaatctg catgcgcaat ttcagcacca gcggaaatct
     1621 cacaagacac atccggaccc ataccggcga gaagcccttt gcctgcgata tctgcggcag
     1681 aaagttcgct ctgtctcagg acctgaaccg gcacaccaag atccacacac accccagagc
     1741 ccctatcccc aagcctttcc agtgccggat ttgcatgcgg aacttcagcg acaggtccgc
     1801 cctgagcaga catatcagaa cacacaccgg cgaaaagcct ttcgcctgtg atatctgtgg
     1861 cagaaagttc gcccggaacg acgacagaaa aaagcacaca aagatccaca ccggatctca
     1921 gaaacctttc caatgtagaa tctgcatgag aaacttcagc cagagcggca acctggccag
     1981 acacatccgg acccacacag gcgaaaagcc tttcgcctgt gatatctgcg gccggaaatt
     2041 cgcccagaga atcgacctga cccggcacac caagatccac ctgagacaga aggacgccgc
     2101 cagaggcagc ggcggcgacg ccaagagcct gaccgcctgg tccaagagga ccgccgacgg
     2161 cagcgagttc gagcccaaga aaaagcggaa ggtgtccggc tctgaaaccc ctggcacaag
     2221 cgagtctgct acacctgagt ccagaaccct ggtcaccttc aaagacgtgt tcgtggactt
     2281 caccagagaa gaatggaagc tgctggatac cgctcagcag atcgtgtaca gaaacgtgat
     2341 gctggaaaac tacaagaatc tggtgtccct gggctaccag ctgaccaagc ctgacgtgat
     2401 cctgagactg gaaaagggcg aggaacccgg gagcggctat ccctatgacg tgcccgatta
     2461 tgcctgatag aattcgctta tcgataatca acctctggat tacaaaattt gtgaaagatt
     2521 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     2581 tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg
     2641 gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac
     2701 tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc
     2761 cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc
     2821 ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa
     2881 atcatcgtcc tttccttggc tgctcgccta tgttgccacc tggattctgc gcgggacgtc
     2941 cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc
     3001 ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg
     3061 ggccgcctcc ccgcatcgat accgagcgct gctcgagaga tctaataaaa tatctttatt
     3121 ttcattacat ctgtgtgttg gttttttgtg tggcggccgc tagtggcagc accagctcag
     3181 ggctggaggt ggtggctcct gagggtacct cagccccagg tggtggcccg ggaaccctgg
     3241 atgacagtgc caccatttgc cgtgtctgcc agaagccagg cgatctggtt atgtgcaacc
     3301 agtgtgagtt ttgtttccac ctggactgtc acctgccggc cctgcaggat gtaccagggg
     3361 aggagtggag ctgctcactc tgccatgtgc tccctgacct gaaggaggag gatggcagcc
     3421 tcagcctgga tggtgcagac agcactggcg tggtggccaa gctctcacca gccaaccagc
     3481 ggaaatgtga gcgtgtactg ctggccctat tctgtcacga accctgccgc cccctgcatc
     3541 agctggctac cgactccacc ttctccctgg accagcccgg tggcaccctg gatctgaccc
     3601 tgatccgtgc ccgcctccag gagaagttgt cacctcccta cagctcccca caggagtttg
     3661 cccaggatgt gggccgcatg ttcaagcaat tcaacaagtt aactgaggac aaggcagacg
     3721 tgcagtccat catcggcctg cagcgcttct tcgagacgcg catgaacgag gccttcggtg
     3781 acaccaagtt ctctgctgtg ctggtggagc ccccgccgat gagcctgcct ggtgctggcc
     3841 tgagttccca ggagctgtct ggtggccctg gtgatggccc ctgcccaact ttaagcttgc
     3901 aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg
     3961 ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc
     4021 gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt tctccttacg catctgtgcg
     4081 gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag
     4141 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc
     4201 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     4261 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     4321 aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     4381 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     4441 actcaactct atctcgggct attcttttga tttataaggg attttgccga tttcggtcta
     4501 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     4561 gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     4621 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     4681 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     4741 atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     4801 catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac
     4861 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     4921 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     4981 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     5041 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     5101 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     5161 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     5221 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     5281 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     5341 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     5401 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     5461 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     5521 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     5581 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     5641 tattgctgat aaatctggag ccggtgagcg tggaagccgc ggtatcattg cagcactggg
     5701 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     5761 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     5821 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     5881 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     5941 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     6001 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     6061 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     6121 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     6181 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     6241 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     6301 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     6361 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     6421 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     6481 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     6541 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     6601 acggttcctg gccttttgct ggccttttgc tcacatgt
//
