LOCUS       sequence-325783-        5969 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Direct-expressing FlpO AAV Virus. Alias: AiP978 - 
            pAAV-AiE2004m-minBG-FlpO-WPRE-HGHpA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5969)
  AUTHORS   Graybuck LT, Daigle TL, Sedeno-Cortes AE, Walker M, Kalmbach B, Lenz
            GH, Morin E, Nguyen TN, Garren E, Bendrick JL, Kim TK, Zhou T, 
            Mortrud M, Yao S, Siverts LA, Larsen R, Gore BB, Szelenyi ER, Trader
            C, Balaram P, van Velthoven CTJ, Chiang M, Mich JK, Dee N, Goldy J, 
            Cetin AH, Smith K, Way SW, Esposito L, Yao Z, Gradinaru V, Sunkin 
            SM, Lein E, Levi BP, Ting JT, Zeng H, Tasic B
  TITLE     Enhancer viruses for combinatorial cell-subclass-specific labeling.
  JOURNAL   Neuron. 2021 Mar 23. pii: S0896-6273(21)00159-8. doi: 
            10.1016/j.neuron.2021.03.011.
  PUBMED    33789083
REFERENCE   2  (bases 1 to 5969)
  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: "Neuron. 
            2021 Mar 23. pii: S0896-6273(21)00159-8. doi: 
            10.1016/j.neuron.2021.03.011."
COMMENT     Sequence Label: AiP978-pAAV-mscRE4-minBGpromoter-FlpO-WPRE-hGHpA
FEATURES             Location/Qualifiers
     source          1..5969
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      767..776
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(773..778,779..799,800..2071)
                     /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"
     misc_feature    2096..2684
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2149..2169)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2567..2578)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2716..3192
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2915..2934)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3232..3372
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3232..3361
                     /label=AAV2 ITR
     rep_origin      3447..3902
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3534..3553)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3744..3765
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3919..3938)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4038..4060
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4098..4116)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4184..4288
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4289..4357,4358..5149)
                     /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(4507..4526)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5320..5908
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5809..5828
                     /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 gcggccgcac gcgtataagc cttgggggca atcaaactat tacattgagt
      181 ccttggattt gctacaaatt acattttaaa tgcaatcatt ttataaaagc ttcaacactc
      241 acacttggaa gcgttaccct gttgaatatc actgactcac taacttgcat tgccatgcta
      301 acttgctttc agagagatct cagaacacat catcttctgc tatttcaata catgcacatt
      361 aatttcctat caacgtgtgc tgatcaggaa ctctgtaatc tggcaccggt gtttattttt
      421 attcctgtct attcctgttg gctcacgaaa agattgtttg agcaagtgtt ttatggtgag
      481 ttgtatcata tgtacattga tttaatctgc ccacattcag ttctacaagc ggagccaaaa
      541 aaatagagac aagcataatt ttcattcaac atgagcccct caatgcaagc caagtacctc
      601 atctggtgct cagctaaagc aacagcaatc tgttccaccc tggagacaca actggccaca
      661 gaaaacttag tgaaaagagg caatgctatg cacaggacaa atgagctcgg gctgggcata
      721 aaagtcaggg cagagccatc tattgcttac atttgcttct ggcgtggcca ccatggctcc
      781 taagaagaag aggaaggtga tgagccagtt cgacatcctg tgcaagaccc cccccaaggt
      841 gctggtgcgg cagttcgtgg agagattcga gaggcccagc ggcgagaaga tcgccagctg
      901 tgccgccgag ctgacctacc tgtgctggat gatcacccac aacggcaccg ccatcaagag
      961 ggccaccttc atgagctaca acaccatcat cagcaacagc ctgagcttcg acatcgtgaa
     1021 caagagcctg cagttcaagt acaagaccca gaaggccacc atcctggagg ccagcctgaa
     1081 gaagctgatc cccgcctggg agttcaccat catcccttac aacggccaga agcaccagag
     1141 cgacatcacc gacatcgtgt ccagcctgca gctgcagttc gagagcagcg aggaggccga
     1201 caagggcaac agccacagca agaagatgct gaaggccctg ctgtccgagg gcgagagcat
     1261 ctgggagatc accgagaaga tcctgaacag cttcgagtac accagcaggt tcaccaagac
     1321 caagaccctg taccagttcc tgttcctggc cacattcatc aactgcggca ggttcagcga
     1381 catcaagaac gtggacccca agagcttcaa gctggtgcag aacaagtacc tgggcgtgat
     1441 cattcagtgc ctggtgaccg agaccaagac aagcgtgtcc aggcacatct actttttcag
     1501 cgccagaggc aggatcgacc ccctggtgta cctggacgag ttcctgagga acagcgagcc
     1561 cgtgctgaag agagtgaaca ggaccggcaa cagcagcagc aacaagcagg agtaccagct
     1621 gctgaaggac aacctggtgc gcagctacaa caaggccctg aagaagaacg ccccctaccc
     1681 catcttcgct atcaagaacg gccctaagag ccacatcggc aggcacctga tgaccagctt
     1741 tctgagcatg aagggcctga ccgagctgac aaacgtggtg ggcaactgga gcgacaagag
     1801 ggcctccgcc gtggccagga ccacctacac ccaccagatc accgccatcc ccgaccacta
     1861 cttcgccctg gtgtccaggt actacgccta cgaccccatc agcaaggaga tgatcgccct
     1921 gaaggacgag accaacccca tcgaggagtg gcagcacatc gagcagctga agggcagcgc
     1981 cgagggcagc atcagatacc ccgcctggaa cggcatcatc agccaggagg tgctggacta
     2041 cctgagcagc tacatcaaca ggcggatctg agaattcgat atcaagctta tcgataatca
     2101 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     2161 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     2221 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc
     2281 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg
     2341 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc
     2401 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     2461 cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgccta
     2521 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc
     2581 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct
     2641 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat accgagcgct
     2701 gctcgagaga tctacgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg
     2761 gaagttgcca ctccagtgcc caccagcctt gtcctaataa aattaagttg catcattttg
     2821 tctgactagg tgtccttcta taatattatg gggtggaggg gggtggtatg gagcaagggg
     2881 caagttggga agacaacctg tagggcctgc ggggtctatt gggaaccaag ctggagtgca
     2941 gtggcacaat cttggctcac tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct
     3001 cagcctcccg agttgttggg attccaggca tgcatgacca ggctcagcta atttttgttt
     3061 ttttggtaga gacggggttt caccatattg gccaggctgg tctccaactc ctaatctcag
     3121 gtgatctacc caccttggcc tcccaaattg ctgggattac aggcgtgaac cactgctccc
     3181 ttccctgtcc ttctgatttt gtaggtaacc acgtgcggac cgagcggccg caggaacccc
     3241 tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac
     3301 caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca
     3361 gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3421 accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg
     3481 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc ccgctccttt
     3541 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     3601 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     3661 tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     3721 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaactc
     3781 tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct attggttaaa
     3841 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat
     3901 tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca
     3961 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     4021 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     4081 acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     4141 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     4201 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     4261 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     4321 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     4381 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     4441 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     4501 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     4561 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     4621 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     4681 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     4741 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     4801 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     4861 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     4921 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     4981 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     5041 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     5101 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     5161 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     5221 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     5281 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     5341 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     5401 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     5461 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     5521 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     5581 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     5641 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     5701 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     5761 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     5821 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     5881 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     5941 ggccttttgc tggccttttg ctcacatgt
//
