LOCUS       sequence-212244-        7275 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector with hSynapsin promoter, Cre-OFF mCAR (for efficient 
            CAV-2 infection) and Cre-ON GCaMP6f (ultrasensitive calcium sensor).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7275)
  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 7275)
  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 {mCAR}off-{GCaMP6f}on-WPRE
FEATURES             Location/Qualifiers
     source          1..7275
                     /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             1845..1874
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(join(1886..2329,2330..2335,2336..2770,2771..2788,
                     2789..3058,3059..3064,3065..3121,3122..3148,3149..3172,
                     3173..3175,3176..3208,3209..3211,3212..3229,3230..3238))
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     primer_bind     2445..2464
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2704..2725
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2831..2852)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      3133..3142
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      3235..3244
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3260..3293
                     /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    3344..3377
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3402..3990
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3455..3475)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3873..3884)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4022..4498
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4221..4240)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4538..4678
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4538..4667
                     /label=AAV2 ITR
     rep_origin      4753..5208
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4840..4859)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5050..5071
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5225..5244)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5344..5366
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5404..5422)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5490..5594
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5595..5663,5664..6455)
                     /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(5813..5832)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6626..7214
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7115..7134
                     /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 ggctcgccta ctgtgcttcg tgctcttgtg cgggatcgcg gatttcacca
      841 gtggtttgag catcactaca cccgaacaga ggatcgaaaa agccaaaggg gaaactgcgt
      901 atctaccatg caagtttact ctcagtcccg aagaccaggg accactggac attgaatggc
      961 tgatatcccc gtctgataac cagatagtgg atcaagtgat cattttgtat tctggagaca
     1021 aaatttatga taactactat ccggatctga aaggacgggt acattttacg agtaacgatg
     1081 tcaagtctgg cgacgcatct ataaatgtga ccaacctgca gctgtcggac attggcactt
     1141 accagtgcaa agtgaagaaa gcccctgggg ttgcaaataa gaaattcctg ctgaccgttc
     1201 ttgttaagcc ttcaggtaca agatgcttcg tggatggatc ggaagagatt ggaaatgact
     1261 tcaagctaaa atgtgaaccc aaggaaggct cccttccact acagtttgaa tggcagaaac
     1321 tgtcggactc ccagacaatg cctacgccat ggctggcaga aatgacgtca ccagttatat
     1381 ctgtgaagaa cgccagttct gagtattctg ggacatacag ctgcacggtt caaaacagag
     1441 tgggctctga ccagtgtatg ctgcgactag acgttgtccc accctccaac cgagccggaa
     1501 cgatcgcggg cgccgtcata gggacgctgc tggcccttgt gctcatcggg gccatcctct
     1561 tctgctgtca caggaaacgc agagaagaga agtacgagaa ggaagttcat catgatatca
     1621 gggaagatgt gcctcctcca aagagtcgga catccactgc caggagctat attggcagca
     1681 accattcctc cctgggatcc atgtccccct ctaacatgga ggggtattcc aagacgcagt
     1741 ataaccaagt ccccagtgaa gactttgaac gtgcgcctca gagcccgact ctggcacccg
     1801 ctaagttcaa gtacgcttac aagaccgatg gcattacagt ggtagagcag aagctgatca
     1861 gcgaggagga cctgtaatta attaattact tcgctgtcat catttgtaca aactcttcgt
     1921 agtttacctg accatcccca tcgatgtctg cttccctgat catttcatca acctcttcat
     1981 ctgttaactt ctctccaagg tttgtcatca cgtggcgaag ctctgctgca ctgatgtagc
     2041 cattgccatc cttatcaaac acaccgaacg cttctctaat ttcttcttcc gtgtccctgt
     2101 atttcatttt tcttgccatc attgtcagga actcagggaa gtcgattgtg ccgtcaccgt
     2161 cggcatctac ttcattgatc atgtcctgca gctctgcttc tgtggggttc tgccccagag
     2221 accgcatcac cgtccccagc tccttggttg ttattgtccc atccccgtcc ttgtcaaata
     2281 gggagaattc ctctttaaat tctgcgatct gctcttcagt cagttggtcc ggcaggttgt
     2341 actccagctt gtgccccagg atgttgccgt cctccttgaa gtcgatgccc ttcagctcga
     2401 tgcggttcac cagggtgtcg ccctcgaact tcacctcggc gcgggtcttg tagttgccgt
     2461 cgtccttgaa gaagatggtg cgctcctgga tgtagccttc gggcatggcg gacttgaaga
     2521 agtcgtgctg cttcatgtgg tcggggtagc ggctgaagca ctgcacgccg taggtcaggg
     2581 tggtcacgag ggtgggccag ggcacgggca gcttgccggt ggtgcagatg aacttcaggg
     2641 tcagcttgcc gtaggtggca tcgccctcac cctcgccgga cacgctgaac ttgtggccgt
     2701 ttacgtcgcc gtccagctcg accaggatgg gcaccacccc ggtgaacagc tcctcgccct
     2761 tgctcaccat gctccctccg gtaccgccct tgtacagctc gtccatgccg agagtgatcc
     2821 cggcggcggt cacgaactcc agcaggacca tgtgatcgcg cttctcgttg gggtctttcg
     2881 aaagtttgga ctgcacgctc aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga
     2941 tgggggtgtt ctgctggtag tggtaggcga gctgcacgcc gccgtcctcg atgttgtggc
     3001 ggatcttgaa gttcgccttg atgccgttct tctgcttgtc ggccttgata tagacgttct
     3061 cgagtgagct cagccgacct atagctctga ctgcgtgacc tgtcttattc cacttacgac
     3121 gtgatgagtc gaccatggtg gcgagatcct tatcgtcatc gtcgtacaga tcccgaccca
     3181 tttgctgtcc accagtcatg ctagccatac catgatgatg atgatgatga gaacccatgg
     3241 tggcaacagc gctactagca taacttcgta taaagtatcc tatacgaagt tatttgcctt
     3301 aacccagaaa ttatcactgt tattctttag aatggtgcaa agaataactt cgtataatgt
     3361 atgctatacg aagttatgaa ttcgatatca agcttatcga taatcaacct ctggattaca
     3421 aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat
     3481 acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct
     3541 ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac
     3601 gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca
     3661 cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca
     3721 tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg
     3781 tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctatgtt gccacctgga
     3841 ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt
     3901 cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga
     3961 gtcggatctc cctttgggcc gcctccccgc atcgataccg agcgctgctc gagagatcta
     4021 cgggtggcat ccctgtgacc cctccccagt gcctctcctg gccctggaag ttgccactcc
     4081 agtgcccacc agccttgtcc taataaaatt aagttgcatc attttgtctg actaggtgtc
     4141 cttctataat attatggggt ggaggggggt ggtatggagc aaggggcaag ttgggaagac
     4201 aacctgtagg gcctgcgggg tctattggga accaagctgg agtgcagtgg cacaatcttg
     4261 gctcactgca atctccgcct cctgggttca agcgattctc ctgcctcagc ctcccgagtt
     4321 gttgggattc caggcatgca tgaccaggct cagctaattt ttgttttttt ggtagagacg
     4381 gggtttcacc atattggcca ggctggtctc caactcctaa tctcaggtga tctacccacc
     4441 ttggcctccc aaattgctgg gattacaggc gtgaaccact gctcccttcc ctgtccttct
     4501 gattttgtag gtaaccacgt gcggaccgag cggccgcagg aacccctagt gatggagttg
     4561 gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga
     4621 cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg
     4681 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa
     4741 agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     4801 gcagcgtgac cgctacactt gccagcgcct tagcgcccgc tcctttcgct ttcttccctt
     4861 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     4921 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt
     4981 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     5041 tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc tcgggctatt
     5101 cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat gagctgattt
     5161 aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc
     5221 tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg
     5281 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
     5341 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa
     5401 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     5461 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     5521 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5581 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     5641 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     5701 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     5761 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     5821 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     5881 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     5941 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     6001 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     6061 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     6121 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     6181 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     6241 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     6301 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     6361 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     6421 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     6481 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     6541 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     6601 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     6661 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     6721 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     6781 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     6841 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     6901 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     6961 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     7021 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     7081 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     7141 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     7201 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     7261 cttttgctca catgt
//
