LOCUS       sequence-466215-        6010 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0064h is an enhancer sequence, designed to drive AAV-mediated 
            transgene expression in specific populations of brain cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6010)
  AUTHORS   Ben-Simon Y, Hooper M, Narayan S, Daigle TL, Dwivedi D, Way SW, 
            Oster A, Stafford DA, Mich JK, Taormina MJ, Martinez RA, Opitz-Araya
            X, Roth JR, Alexander JR, Allen S, Amster A, Arbuckle J, Ayala A, 
            Baker PM, Bakken TE, Barcelli T, Barta S, Bendrick J, Bertagnolli D,
            Bielstein C, Bishwakarma P, Bowlus J, Boyer G, Brouner K, Casian B, 
            Casper T, Chakka AB, Chakrabarty R, Chance RK, Chavan S, Clark M, 
            Colbert K, Collman F, Daniel S, Departee M, DiValentin P, Donadio N,
            Dotson N, Egdorf T, Fliss T, Gabitto M, Garcia J, Gary A, Gasperini 
            M, Gloe J, Goldy J, Gore BB, Graybuck L, Greisman N, Haeseleer F, 
            Halterman C, Haradon Z, Hastings SD, Helback O, Ho W, Hockemeyer D, 
            Huang C, Huff S, Hunker A, Johansen N, Jones D, Juneau Z, Kalmbach 
            B, Kannan M, Khem S, Kussick E, Kutsal R, Larsen R, Lee C, Lee AY, 
            Leibly M, Lenz GH, Li S, Liang E, Lusk N, Madigan Z, Malloy J, 
            Malone J, McCue R, Melchor J, Mollenkopf T, Moosman S, Morin E, 
            Newman D, Ng L, Ngo K, Omstead V, Otto S, Oyama A, Pena N, Pham T, 
            Phillips E, Pom CA, Potekhina L, Ransford S, Ray PL, Rette D, 
            Reynoldson C, Rimorin C, Rocha D, Ruiz A, Sanchez REA, Sawyer L, 
            Sedeno-Cortes A, Sevigny JP, Shapovalova N, Shepard N, Shulga L, 
            Sigler AR, Siverts L, Soliman S, Somasundaram S, Staats B, Stewart 
            K, Szelenyi E, Tieu M, Trader C, Tran A, van Velthoven CTJ, Walker 
            M, Wang Y, Weed N, Wirthlin M, Wood T, Wynalda B, Yao Z, Zhou T, 
            Ariza J, Dee N, Reding M, Ronellenfitch K, Mufti S, Sunkin SM, Smith
            KA, Esposito L, Waters J, Thyagarajan B, Yao S, Lein ES, Zeng H, 
            Levi BP, Ngai J, Ting JT, Tasic B
  TITLE     A suite of enhancer AAVs and transgenic mouse lines for genetic 
            access to cortical cell types.
  JOURNAL   Cell. 2025 May 16:S0092-8674(25)00513-6. doi: 
            10.1016/j.cell.2025.05.002.
  PUBMED    40403729
REFERENCE   2  (bases 1 to 6010)
  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: "Cell. 2025 
            May 16:S0092-8674(25)00513-6. doi: 10.1016/j.cell.2025.05.002."
COMMENT     Sequence Label: AiP1224 - pAAV-AiE0064h-minBG-FlpO-WPRE-HGHpA
FEATURES             Location/Qualifiers
     source          1..6010
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      180..189
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     repeat_region   867..996
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(1058..1646)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1138..1157)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1817..2608,2609..2677))
                     /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     2440..2459
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2678..2782)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2850..2868
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2906..2928)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3028..3047
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(3064..3519)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3201..3222)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3413..3432
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   3594..3734
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3605..3734
                     /label=AAV2 ITR
     polyA_signal    3774..4250
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     4032..4051
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     misc_feature    4282..4870
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             4388..4399
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     4797..4817
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(4895..6010,1..156,157..177,178..183))
                     /codon_start=1
                     /product="nuclear-targeted site-specific recombinase"
                     /label=FLPo
                     /note="FLPo is a mouse codon-optimized fusion between the 
                     SV40 nuclear localization signal and a thermostable version
                     of the Saccharomyces cerevisiae site-specific recombinase 
                     FLP.  Recombination occurs at FRT sequences."
                     /translation="MAPKKKRKVMSQFDILCKTPPKVLVRQFVERFERPSGEKIASCAA
                     ELTYLCWMITHNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKK
                     LIPAWEFTIIPYNGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESI
                     WEITEKILNSFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGV
                     IIQCLVTETKTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEY
                     QLLKDNLVRSYNKALKKNAPYPIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWS
                     DKRASAVARTTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQL
                     KGSAEGSIRYPAWNGIISQEVLDYLSSYINRRI"
ORIGIN
        1 gatggcggtg ccgttgtggg tgatcatcca gcacaggtag gtcagctcgg cggcacagct
       61 ggcgatcttc tcgccgctgg gcctctcgaa tctctccacg aactgccgca ccagcacctt
      121 ggggggggtc ttgcacagga tgtcgaactg gctcatcacc ttcctcttct tcttaggagc
      181 catggtggcc acgccagaag caaatgtaag caatagatgg ctctgccctg acttttatgc
      241 ccagcccgag ctcgcaaata catgagttat cttaacctag tcatgtgaga aagtagaagg
      301 aaatgaggat gactcagagg gaaaatgcta ccagtgcaag aaaatgcggt caaactcagc
      361 atgaatttta aggtccatac agaagtttat tcaaaaacaa gttgataggg gttcttggct
      421 aatatttttt aggcaaaatg tattgagtta aatgtcatgc caaatacttt tcaaagcttt
      481 actgtatttt gcaatctaaa aatccaattt tgtattttat gctacagtta gcactccttc
      541 acaacgcagt tctgtcaatg agatacttcc ttcatgtcaa aagaaaattc caataatcag
      601 aaaaacagct gtgtagatga ctcacactac gttctgctag gagagaattt ttgtaaaaga
      661 ggcaagtaca agagagttca attgtaatta tataccacaa ctttaccaat tgaaaacact
      721 tcttacatca cagggaaagt attcaagtag tcagaagggg cacctgagta tgcaacttgc
      781 cgcataactg aaatagaaga gggggttagt caaagcagct agggcaaaca catttgagat
      841 ttaagagttc atacgcgtgc ggccgcagga acccctagtg atggagttgg ccactccctc
      901 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggcgg
      961 cctcagtgag cgagcgagcg cgcagctgcc tgcaggacat gtgagcaaaa ggccagcaaa
     1021 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     1081 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     1141 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     1201 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     1261 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     1321 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     1381 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     1441 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     1501 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     1561 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     1621 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     1681 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     1741 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     1801 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     1861 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     1921 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     1981 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     2041 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     2101 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     2161 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     2221 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     2281 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     2341 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     2401 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     2461 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     2521 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     2581 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     2641 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     2701 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     2761 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     2821 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtctcg
     2881 cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag
     2941 cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg
     3001 gcgggtgtcg gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc
     3061 ataaaattgt aaacgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     3121 cattttttaa ccaatagacc gaaatcggca aaatccctta taaatcaaaa gaatagcccg
     3181 agatagagtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     3241 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     3301 ccaaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     3361 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     3421 aagcgaaagg agcgggcgct aaggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     3481 ccacacccgc cgcgcttaat gcgccgctac agggcgcgta ctatggttgc tttgacgtat
     3541 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gcccctgcag
     3601 gcagctgcgc gctcgctcgc tcactgaggc cgcccgggca aagcccgggc gtcgggcgac
     3661 ctttggtcgc ccggcctcag tgagcgagcg agcgcgcaga gagggagtgg ccaactccat
     3721 cactaggggt tcctgcggcc gctcggtccg cacgtggtta cctacaaaat cagaaggaca
     3781 gggaagggag cagtggttca cgcctgtaat cccagcaatt tgggaggcca aggtgggtag
     3841 atcacctgag attaggagtt ggagaccagc ctggccaata tggtgaaacc ccgtctctac
     3901 caaaaaaaca aaaattagct gagcctggtc atgcatgcct ggaatcccaa caactcggga
     3961 ggctgaggca ggagaatcgc ttgaacccag gaggcggaga ttgcagtgag ccaagattgt
     4021 gccactgcac tccagcttgg ttcccaatag accccgcagg ccctacaggt tgtcttccca
     4081 acttgcccct tgctccatac cacccccctc caccccataa tattatagaa ggacacctag
     4141 tcagacaaaa tgatgcaact taattttatt aggacaaggc tggtgggcac tggagtggca
     4201 acttccaggg ccaggagagg cactggggag gggtcacagg gatgccaccc gtagatctct
     4261 cgagcagcgc tcggtatcga tgcggggagg cggcccaaag ggagatccga ctcgtctgag
     4321 ggcgaaggcg aagacgcgga agaggccgca gagccggcag caggccgcgg gaaggaaggt
     4381 ccgctggatt gagggccgaa gggacgtagc agaaggacgt cccgcgcaga atccaggtgg
     4441 caacataggc gagcagccaa ggaaaggacg atgatttccc cgacaacacc acggaattgt
     4501 cagtgcccaa cagccgagcc cctgtccagc agcgggcaag gcaggcggcg atgagttccg
     4561 ccgtggcaat agggaggggg aaagcgaaag tcccggaaag gagctgacag gtggtggcaa
     4621 tgccccaacc agtgggggtt gcgtcagcaa acacagtgca caccacgcca cgttgcctga
     4681 caacgggcca caactcctca taaagagaca gcaaccagga tttatacaag gaggagaaaa
     4741 tgaaagccat acgggaagca atagcatgat acaaaggcat taaagcagcg tatccacata
     4801 gcgtaaaagg agcaacatag ttaagaatac cagtcaatct ttcacaaatt ttgtaatcca
     4861 gaggttgatt atcgataagc ttgatatcga attctcagat ccgcctgttg atgtagctgc
     4921 tcaggtagtc cagcacctcc tggctgatga tgccgttcca ggcggggtat ctgatgctgc
     4981 cctcggcgct gcccttcagc tgctcgatgt gctgccactc ctcgatgggg ttggtctcgt
     5041 ccttcagggc gatcatctcc ttgctgatgg ggtcgtaggc gtagtacctg gacaccaggg
     5101 cgaagtagtg gtcggggatg gcggtgatct ggtgggtgta ggtggtcctg gccacggcgg
     5161 aggccctctt gtcgctccag ttgcccacca cgtttgtcag ctcggtcagg cccttcatgc
     5221 tcagaaagct ggtcatcagg tgcctgccga tgtggctctt agggccgttc ttgatagcga
     5281 agatggggta gggggcgttc ttcttcaggg ccttgttgta gctgcgcacc aggttgtcct
     5341 tcagcagctg gtactcctgc ttgttgctgc tgctgttgcc ggtcctgttc actctcttca
     5401 gcacgggctc gctgttcctc aggaactcgt ccaggtacac cagggggtcg atcctgcctc
     5461 tggcgctgaa aaagtagatg tgcctggaca cgcttgtctt ggtctcggtc accaggcact
     5521 gaatgatcac gcccaggtac ttgttctgca ccagcttgaa gctcttgggg tccacgttct
     5581 tgatgtcgct gaacctgccg cagttgatga atgtggccag gaacaggaac tggtacaggg
     5641 tcttggtctt ggtgaacctg ctggtgtact cgaagctgtt caggatcttc tcggtgatct
     5701 cccagatgct ctcgccctcg gacagcaggg ccttcagcat cttcttgctg tggctgttgc
     5761 ccttgtcggc ctcctcgctg ctctcgaact gcagctgcag gctggacacg atgtcggtga
     5821 tgtcgctctg gtgcttctgg ccgttgtaag ggatgatggt gaactcccag gcggggatca
     5881 gcttcttcag gctggcctcc aggatggtgg ccttctgggt cttgtacttg aactgcaggc
     5941 tcttgttcac gatgtcgaag ctcaggctgt tgctgatgat ggtgttgtag ctcatgaagg
     6001 tggccctctt
//
