LOCUS       sequence-209262-        6616 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector with hSynapsin promoter, Cre-OFF hCAR (for efficient 
            CAV-2 infection), Cre-ON DTR-EGFP (diphtheria toxin receptor for 
            cell ablation), and W3SL cassette (for maximize cloning capacity).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6616)
  AUTHORS   Li SJ, Vaughan A, Sturgill JF, Kepecs A
  TITLE     A Viral Receptor Complementation Strategy to Overcome CAV-2 Tropism 
            for Efficient Retrograde Targeting of Neurons.
  JOURNAL   Neuron. 2018 Jun 6;98(5):905-917.e5. doi: 
            10.1016/j.neuron.2018.05.028.
  PUBMED    29879392
REFERENCE   2  (bases 1 to 6616)
  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.neuron.2018.05"; journalName: "Neuron"; date: "2018-06-6-
            6"; volume: "98"; issue: "5"; pages: "905-917.e5"
COMMENT     Sequence Label: pAAV-hSyn-DIO {hCAR}off-{DTR-GFP}on-W3SL
FEATURES             Location/Qualifiers
     source          1..6616
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        186..633
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(671..704)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(755..788)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      794..803
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1895..1924
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(join(1936..2649,2650..2652,2653..2655))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(1939..2649,2650..2652,2653..2655))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1981..2002)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2330..2349
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2589..2610
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2652..2661
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3311..3344
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    3395..3428
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(3498..3518)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3878..4018
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3878..4007
                     /label=AAV2 ITR
     rep_origin      4093..4548
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4180..4199)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4390..4411
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4565..4584)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4684..4706
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4744..4762)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4830..4934
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4935..5003,5004..5795)
                     /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(5153..5172)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5966..6554
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6455..6474
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtgtgtc tagactgcag agggccctgc
      181 gtatgagtgc aagtgggttt taggaccagg atgaggcggg gtgggggtgc ctacctgacg
      241 accgaccccg acccactgga caagcaccca acccccattc cccaaattgc gcatccccta
      301 tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt gcgcactgcc agcttcagca
      361 ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc accgccgcct cagcactgaa
      421 ggcgcgctga cgtcactcgc cggtcccccg caaactcccc ttcccggcca ccttggtcgc
      481 gtccgcgccg ccgccggccc agccggaccg caccacgcga ggcgcgagat aggggggcac
      541 gggcgcgacc atctgcgctg cggcgccggc gactcagcgc tgcctcagtc tgcggtgggc
      601 agcggaggag tcgtgtcgtg cctgagagcg cagtcgagaa ggtaccggat cctctagagt
      661 cgactccgga ataacttcgt ataggatact ttatacgaag ttatgcagaa tggtagctgg
      721 attgtagctg ctattagcaa tatgaaacct cttaataact tcgtatagca tacattatac
      781 gaagttatgg cgcgccacca tggcgctcct gctgtgcttc gtgctcctgt gcggagtagt
      841 ggatttcgcc agaagtttga gtatcactac tcctgaagag atgattgaaa aagccaaagg
      901 ggaaactgcc tatctgccgt gcaaatttac gcttagtccc gaagaccagg gaccgctgga
      961 catcgagtgg ctgatatcac cagctgataa tcagaaggtg gatcaagtga ttattttata
     1021 ttctggagac aaaatttatg atgactacta tccagatctg aaaggccgag tacattttac
     1081 gagtaatgat ctcaaatctg gtgatgcatc aataaatgta acgaatttac aactgtcaga
     1141 tattggcaca tatcagtgca aagtgaaaaa agctcctggt gttgcaaata agaagattca
     1201 tctggtagtt cttgttaagc cttcaggtgc gagatgttac gttgatggat ctgaagaaat
     1261 tggaagtgac tttaagataa aatgtgaacc aaaagaaggt tcacttccat tacagtatga
     1321 gtggcaaaaa ttgtctgact cacagaaaat gcccacttca tggttagcag aaatgacttc
     1381 atctgttata tctgtaaaaa atgcctcttc tgagtactct gggacataca gctgtacagt
     1441 cagaaacaga gtgggctctg atcagtgcct gttgcgtcta aacgttgtcc ctccttcaaa
     1501 taaagctgga ctaattgcag gagccattat aggaactttg cttgctctag cgctcattgg
     1561 tcttatcatc ttttgctgtc gtaaaaagcg cagagaagaa aaatatgaaa aggaagttca
     1621 tcacgatatc agggaagatg tgccacctcc aaagagccgt acgtccactg ccagaagcta
     1681 catcggcagt aatcattcat ccctggggtc catgtctcct tccaacatgg aaggatattc
     1741 caagactcag tataaccaag taccaagtga agactttgaa cgcactcctc agagtccgac
     1801 tctcccacct gctaaggtag ctgcccctaa tctaagtcga atgggtgcga ttcctgtgat
     1861 gattccagca cagagcaagg atgggtctat agtagagcag aagctgatca gcgaggagga
     1921 cctgtaatta attaattact tgtacagctc gtccatgccg agagtgatcc cggcggcggt
     1981 cacgaactcc agcaggacca tgtgatcgcg cttctcgttg gggtctttgc tcagggcgga
     2041 ctgggtgctc aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt
     2101 ctgctggtag tggtcggcga gctgcacgct gccgtcctcg atgttgtggc ggatcttgaa
     2161 gttcaccttg atgccgttct tctgcttgtc ggccatgata tagacgttgt ggctgttgta
     2221 gttgtactcc agcttgtgcc ccaggatgtt gccgtcctcc ttgaagtcga tgcccttcag
     2281 ctcgatgcgg ttcaccaggg tgtcgccctc gaacttcacc tcggcgcggg tcttgtagtt
     2341 gccgtcgtcc ttgaagaaga tggtgcgctc ctggacgtag ccttcgggca tggcggactt
     2401 gaagaagtcg tgctgcttca tgtggtcggg gtagcggctg aagcactgca cgccgtaggt
     2461 cagggtggtc acgagggtgg gccagggcac gggcagcttg ccggtggtgc agatgaactt
     2521 cagggtcagc ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg
     2581 gccgtttacg tcgccgtcca gctcgaccag gatgggcacc accccggtga acagctcctc
     2641 gcccttgctc accatggtgg cgaccggtgg atccgggtgg gaattagtca tgcccaactt
     2701 cactttctct tcgttttcca catcataacc acctctccta tggtacctaa acatgagaag
     2761 ccccacgatg accagcagac agacagagga cagcaccacg gccaccacag ccaggatagt
     2821 tgtatggtca taggtatata agcgattttc cactgggagg ctcagcccat gacacctctc
     2881 tccatggtaa cctgggtggc agatgcagga gggagcccgg agctccttca catatttgca
     2941 ttctccgtgg atgcagaagt ccttgtattt ccgaagacat gggtccctct tcttccctag
     3001 tcccttgcct ttcttctttc ttttcccgtg ctcctccttg cttggtgtgg ccagtgcttg
     3061 tggcttggag gataaagtga ctctcaaaag gtccagatct gcctcttgca agtcacggac
     3121 tttccggtcc cggccgcctc ctaggcgtag cagctggtcc gtagatccag tggaagggtc
     3181 cgggttgctg gttccagcag ctagccctct ccgaagctgc tccaggctct cgccagtcac
     3241 cagtgccgaa agaactgcag ccagaaggag cttcagcacc accgacggca gcagcttcat
     3301 ggtgactagc ataacttcgt ataaagtatc ctatacgaag ttatttgcct taacccagaa
     3361 attatcactg ttattcttta gaatggtgca aagaataact tcgtataatg tatgctatac
     3421 gaagttatga attccgctcg agataatcaa cctctggatt acaaaatttg tgaaagattg
     3481 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     3541 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     3601 ttagttcttg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     3661 cggctgttgg gcactgacaa ttccgtggtg tttatttgtg aaatttgtga tgctattgct
     3721 ttatttgtaa ccatctagct ttatttgtga aatttgtgat gctattgctt tatttgtaac
     3781 cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt
     3841 tcagggggag atgtgggagg ttttttaaag cggccgcagg aacccctagt gatggagttg
     3901 gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga
     3961 cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg
     4021 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa
     4081 agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     4141 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     4201 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     4261 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt
     4321 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     4381 tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc tcgggctatt
     4441 cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat gagctgattt
     4501 aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc
     4561 tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg
     4621 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
     4681 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa
     4741 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     4801 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     4861 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     4921 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     4981 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     5041 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     5101 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     5161 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     5221 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     5281 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     5341 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     5401 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     5461 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     5521 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     5581 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     5641 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     5701 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     5761 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     5821 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     5881 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     5941 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     6001 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     6061 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     6121 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     6181 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     6241 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     6301 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     6361 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     6421 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     6481 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     6541 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     6601 cttttgctca catgtt
//
