LOCUS       sequence-212262-        6604 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 6604)
  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 6604)
  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..6604
                     /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        175..622
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(660..693)
                     /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(744..777)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      783..792
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1884..1913
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(join(1925..2638,2639..2641,2642..2644))
                     /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(1928..2638,2639..2641,2642..2644))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1970..1991)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2319..2338
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2578..2599
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2641..2650
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3300..3333
                     /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    3384..3417
                     /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(3487..3507)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3867..4007
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3867..3996
                     /label=AAV2 ITR
     rep_origin      4082..4537
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4169..4188)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4379..4400
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4554..4573)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4673..4695
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4733..4751)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4819..4923
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4924..4992,4993..5784)
                     /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(5142..5161)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5955..6543
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6444..6463
                     /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 gcgtgtgtct agactgcaga gggccctgcg tatgagtgca
      181 agtgggtttt aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga
      241 cccactggac aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg
      301 gaggggaaac aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt
      361 gccttcgccc ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac
      421 gtcactcgcc ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc
      481 cgccggccca gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca
      541 tctgcgctgc ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt
      601 cgtgtcgtgc ctgagagcgc agtcgagaag gtaccggatc ctctagagtc gactccggaa
      661 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
      721 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
      781 gcgccaccat ggcgctcctg ctgtgcttcg tgctcctgtg cggagtagtg gatttcgcca
      841 gaagtttgag tatcactact cctgaagaga tgattgaaaa agccaaaggg gaaactgcct
      901 atctgccgtg caaatttacg cttagtcccg aagaccaggg accgctggac atcgagtggc
      961 tgatatcacc agctgataat cagaaggtgg atcaagtgat tattttatat tctggagaca
     1021 aaatttatga tgactactat ccagatctga aaggccgagt acattttacg agtaatgatc
     1081 tcaaatctgg tgatgcatca ataaatgtaa cgaatttaca actgtcagat attggcacat
     1141 atcagtgcaa agtgaaaaaa gctcctggtg ttgcaaataa gaagattcat ctggtagttc
     1201 ttgttaagcc ttcaggtgcg agatgttacg ttgatggatc tgaagaaatt ggaagtgact
     1261 ttaagataaa atgtgaacca aaagaaggtt cacttccatt acagtatgag tggcaaaaat
     1321 tgtctgactc acagaaaatg cccacttcat ggttagcaga aatgacttca tctgttatat
     1381 ctgtaaaaaa tgcctcttct gagtactctg ggacatacag ctgtacagtc agaaacagag
     1441 tgggctctga tcagtgcctg ttgcgtctaa acgttgtccc tccttcaaat aaagctggac
     1501 taattgcagg agccattata ggaactttgc ttgctctagc gctcattggt cttatcatct
     1561 tttgctgtcg taaaaagcgc agagaagaaa aatatgaaaa ggaagttcat cacgatatca
     1621 gggaagatgt gccacctcca aagagccgta cgtccactgc cagaagctac atcggcagta
     1681 atcattcatc cctggggtcc atgtctcctt ccaacatgga aggatattcc aagactcagt
     1741 ataaccaagt accaagtgaa gactttgaac gcactcctca gagtccgact ctcccacctg
     1801 ctaaggtagc tgcccctaat ctaagtcgaa tgggtgcgat tcctgtgatg attccagcac
     1861 agagcaagga tgggtctata gtagagcaga agctgatcag cgaggaggac ctgtaattaa
     1921 ttaattactt gtacagctcg tccatgccga gagtgatccc ggcggcggtc acgaactcca
     1981 gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct cagggcggac tgggtgctca
     2041 ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat gggggtgttc tgctggtagt
     2101 ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggcg gatcttgaag ttcaccttga
     2161 tgccgttctt ctgcttgtcg gccatgatat agacgttgtg gctgttgtag ttgtactcca
     2221 gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat gcccttcagc tcgatgcggt
     2281 tcaccagggt gtcgccctcg aacttcacct cggcgcgggt cttgtagttg ccgtcgtcct
     2341 tgaagaagat ggtgcgctcc tggacgtagc cttcgggcat ggcggacttg aagaagtcgt
     2401 gctgcttcat gtggtcgggg tagcggctga agcactgcac gccgtaggtc agggtggtca
     2461 cgagggtggg ccagggcacg ggcagcttgc cggtggtgca gatgaacttc agggtcagct
     2521 tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct gaacttgtgg ccgtttacgt
     2581 cgccgtccag ctcgaccagg atgggcacca ccccggtgaa cagctcctcg cccttgctca
     2641 ccatggtggc gaccggtgga tccgggtggg aattagtcat gcccaacttc actttctctt
     2701 cgttttccac atcataacca cctctcctat ggtacctaaa catgagaagc cccacgatga
     2761 ccagcagaca gacagaggac agcaccacgg ccaccacagc caggatagtt gtatggtcat
     2821 aggtatataa gcgattttcc actgggaggc tcagcccatg acacctctct ccatggtaac
     2881 ctgggtggca gatgcaggag ggagcccgga gctccttcac atatttgcat tctccgtgga
     2941 tgcagaagtc cttgtatttc cgaagacatg ggtccctctt cttccctagt cccttgcctt
     3001 tcttctttct tttcccgtgc tcctccttgc ttggtgtggc cagtgcttgt ggcttggagg
     3061 ataaagtgac tctcaaaagg tccagatctg cctcttgcaa gtcacggact ttccggtccc
     3121 ggccgcctcc taggcgtagc agctggtccg tagatccagt ggaagggtcc gggttgctgg
     3181 ttccagcagc tagccctctc cgaagctgct ccaggctctc gccagtcacc agtgccgaaa
     3241 gaactgcagc cagaaggagc ttcagcacca ccgacggcag cagcttcatg gtgactagca
     3301 taacttcgta taaagtatcc tatacgaagt tatttgcctt aacccagaaa ttatcactgt
     3361 tattctttag aatggtgcaa agaataactt cgtataatgt atgctatacg aagttatgaa
     3421 ttccgctcga gataatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     3481 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     3541 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tagttcttgc
     3601 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     3661 cactgacaat tccgtggtgt ttatttgtga aatttgtgat gctattgctt tatttgtaac
     3721 catctagctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     3781 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggaga
     3841 tgtgggaggt tttttaaagc ggccgcagga acccctagtg atggagttgg ccactccctc
     3901 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt
     3961 tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg
     4021 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag
     4081 tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     4141 gctacacttg ccagcgcctt agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     4201 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     4261 agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg
     4321 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     4381 ggactcttgt tccaaactgg aacaacactc aactctatct cgggctattc ttttgattta
     4441 taagggattt tgccgatttc ggtctattgg ttaaaaaatg agctgattta acaaaaattt
     4501 aacgcgaatt ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc
     4561 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc
     4621 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
     4681 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg
     4741 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
     4801 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     4861 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     4921 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     4981 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     5041 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     5101 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     5161 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     5221 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     5281 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     5341 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     5401 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     5461 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     5521 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     5581 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     5641 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     5701 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     5761 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     5821 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     5881 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     5941 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     6001 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     6061 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     6121 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     6181 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     6241 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     6301 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     6361 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     6421 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     6481 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     6541 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     6601 atgt
//
