LOCUS       sequence-341012-        5896 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV expression of Flpo-3xFLAG by hSyn1 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5896)
  AUTHORS   Kitanishi T, Tashiro M, Kitanishi N, Mizuseki K
  TITLE     Intersectional, anterograde transsynaptic targeting of neurons 
            receiving monosynaptic inputs from two upstream regions.
  JOURNAL   Commun Biol. 2022 Feb 21;5(1):149. doi: 10.1038/s42003-022-03096-3.
  PUBMED    35190665
REFERENCE   2  (bases 1 to 5896)
  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: "Commun 
            Biol."; date: "2022-02-21"; volume: "5(1):149. doi"; pages: " 
            10.1038/s42003-022-03096-3"
COMMENT     Sequence Label: pAAV-hSyn-Flpo-3xFLAG
FEATURES             Location/Qualifiers
     source          1..5896
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   21..150
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        195..642
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      656..665
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1952..2017
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     misc_feature    2043..2631
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2096..2116)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2514..2525)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2663..3139
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2862..2881)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3179..3319
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3179..3308
                     /label=AAV2 ITR
     rep_origin      3394..3849
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3481..3500)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3691..3712
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3866..3885)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3985..4007
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4045..4063)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4131..4235
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4236..4304,4305..5096)
                     /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(4454..4473)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5267..5855
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5756..5775
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctggcctttt gctcacatgt cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc
       61 ccgggcgtcg ggcgaccttt ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg
      121 gagtggccaa ctccatcact aggggttcct gcggccgcac gcgtgtgtct agactgcaga
      181 gggccctgcg tatgagtgca agtgggtttt aggaccagga tgaggcgggg tgggggtgcc
      241 tacctgacga ccgaccccga cccactggac aagcacccaa cccccattcc ccaaattgcg
      301 catcccctat cagagagggg gaggggaaac aggatgcggc gaggcgcgtg cgcactgcca
      361 gcttcagcac cgcggacagt gccttcgccc ccgcctggcg gcgcgcgcca ccgccgcctc
      421 agcactgaag gcgcgctgac gtcactcgcc ggtcccccgc aaactcccct tcccggccac
      481 cttggtcgcg tccgcgccgc cgccggccca gccggaccgc accacgcgag gcgcgagata
      541 ggggggcacg ggcgcgacca tctgcgctgc ggcgccggcg actcagcgct gcctcagtct
      601 gcggtgggca gcggaggagt cgtgtcgtgc ctgagagcgc agtcgagaag gtaccgccac
      661 catggccaca accatgagcc agttcgacat cctgtgcaag acccccccca aggtgctggt
      721 gcggcagttc gtggagagat tcgagaggcc cagcggcgag aagatcgcca gctgtgccgc
      781 cgagctgacc tacctgtgct ggatgatcac ccacaacggc accgccatca agagggccac
      841 cttcatgagc tacaacacca tcatcagcaa cagcctgagc ttcgacatcg tgaacaagag
      901 cctgcagttc aagtacaaga cccagaaggc caccatcctg gaggccagcc tgaagaagct
      961 gatccccgcc tgggagttca ccatcatccc ttacaacggc cagaagcacc agagcgacat
     1021 caccgacatc gtgtccagcc tgcagctgca gttcgagagc agcgaggagg ccgacaaggg
     1081 caacagccac agcaagaaga tgctgaaggc cctgctgtcc gagggcgaga gcatctggga
     1141 gatcaccgag aagatcctga acagcttcga gtacaccagc aggttcacca agaccaagac
     1201 cctgtaccag ttcctgttcc tggccacatt catcaactgc ggcaggttca gcgacatcaa
     1261 gaacgtggac cccaagagct tcaagctggt gcagaacaag tacctgggcg tgatcattca
     1321 gtgcctggtg accgagacca agacaagcgt gtccaggcac atctactttt tcagcgccag
     1381 aggcaggatc gaccccctgg tgtacctgga cgagttcctg aggaacagcg agcccgtgct
     1441 gaagagagtg aacaggaccg gcaacagcag cagcaacaag caggagtacc agctgctgaa
     1501 ggacaacctg gtgcgcagct acaacaaggc cctgaagaag aacgccccct accccatctt
     1561 cgctatcaag aacggcccta agagccacat cggcaggcac ctgatgacca gctttctgag
     1621 catgaagggc ctgaccgagc tgacaaacgt ggtgggcaac tggagcgaca agagggcctc
     1681 cgccgtggcc aggaccacct acacccacca gatcaccgcc atccccgacc actacttcgc
     1741 cctggtgtcc aggtactacg cctacgaccc catcagcaag gagatgatcg ccctgaagga
     1801 cgagaccaac cccatcgagg agtggcagca catcgagcag ctgaagggca gcgccgaggg
     1861 cagcatcaga taccccgcct ggaacggcat catcagccag gaggtgctgg actacctgag
     1921 cagctacatc aacaggcgga tcctcgagta cgactacaaa gaccatgacg gtgattataa
     1981 agatcatgac atcgactaca aggatgacga tgacaagtag cgctatcgat aagcttatcg
     2041 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     2101 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     2161 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
     2221 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
     2281 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
     2341 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
     2401 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
     2461 tcgcctatgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
     2521 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
     2581 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc
     2641 gagcgctgct cgagagatct acgggtggca tccctgtgac ccctccccag tgcctctcct
     2701 ggccctggaa gttgccactc cagtgcccac cagccttgtc ctaataaaat taagttgcat
     2761 cattttgtct gactaggtgt ccttctataa tattatgggg tggagggggg tggtatggag
     2821 caaggggcaa gttgggaaga caacctgtag ggcctgcggg gtctattggg aaccaagctg
     2881 gagtgcagtg gcacaatctt ggctcactgc aatctccgcc tcctgggttc aagcgattct
     2941 cctgcctcag cctcccgagt tgttgggatt ccaggcatgc atgaccaggc tcagctaatt
     3001 tttgtttttt tggtagagac ggggtttcac catattggcc aggctggtct ccaactccta
     3061 atctcaggtg atctacccac cttggcctcc caaattgctg ggattacagg cgtgaaccac
     3121 tgctcccttc cctgtccttc tgattttgta ggtaaccacg tgcggaccga gcggccgcag
     3181 gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc
     3241 gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga
     3301 gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     3361 atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg
     3421 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg
     3481 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     3541 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     3601 aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     3661 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     3721 tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt tcggtctatt
     3781 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt
     3841 ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     3901 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     3961 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     4021 caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     4081 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     4141 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     4201 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     4261 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     4321 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     4381 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     4441 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     4501 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     4561 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     4621 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     4681 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     4741 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     4801 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     4861 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     4921 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     4981 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     5041 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     5101 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     5161 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     5221 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     5281 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     5341 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     5401 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     5461 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     5521 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     5581 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     5641 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     5701 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     5761 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     5821 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     5881 ggttcctggc cttttg
//
