LOCUS       sequence-212394-        6603 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector with hSynapsin promoter, Cre-OFF hCAR (for efficient 
            CAV-2 infection), Cre-ON GCaMP6f (ultrasensitive calcium sensor), 
            and W3SL regulatory cassette (for maximize cloning capacity).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6603)
  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 6603)
  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-{GCaMP6f}on-W3SL
FEATURES             Location/Qualifiers
     source          1..6603
                     /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..2368,2369..2374,2375..2809,2810..2827,
                     2828..3097,3098..3103,3104..3160,3161..3187,3188..3211,
                     3212..3214,3215..3247,3248..3250,3251..3268,3269..3277))
                     /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     2484..2503
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2743..2764
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2870..2891)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      3172..3181
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      3274..3283
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3299..3332
                     /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    3383..3416
                     /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(3486..3506)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3866..4006
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3866..3995
                     /label=AAV2 ITR
     rep_origin      4081..4536
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4168..4187)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4378..4399
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4553..4572)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4672..4694
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4732..4750)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4818..4922
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4923..4991,4992..5783)
                     /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(5141..5160)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5954..6542
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6443..6462
                     /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 cgctgtcatc atttgtacaa actcttcgta gtttacctga ccatccccat
     1981 cgatgtctgc ttccctgatc atttcatcaa cctcttcatc tgttaacttc tctccaaggt
     2041 ttgtcatcac gtggcgaagc tctgctgcac tgatgtagcc attgccatcc ttatcaaaca
     2101 caccgaacgc ttctctaatt tcttcttccg tgtccctgta tttcattttt cttgccatca
     2161 ttgtcaggaa ctcagggaag tcgattgtgc cgtcaccgtc ggcatctact tcattgatca
     2221 tgtcctgcag ctctgcttct gtggggttct gccccagaga ccgcatcacc gtccccagct
     2281 ccttggttgt tattgtccca tccccgtcct tgtcaaatag ggagaattcc tctttaaatt
     2341 ctgcgatctg ctcttcagtc agttggtccg gcaggttgta ctccagcttg tgccccagga
     2401 tgttgccgtc ctccttgaag tcgatgccct tcagctcgat gcggttcacc agggtgtcgc
     2461 cctcgaactt cacctcggcg cgggtcttgt agttgccgtc gtccttgaag aagatggtgc
     2521 gctcctggat gtagccttcg ggcatggcgg acttgaagaa gtcgtgctgc ttcatgtggt
     2581 cggggtagcg gctgaagcac tgcacgccgt aggtcagggt ggtcacgagg gtgggccagg
     2641 gcacgggcag cttgccggtg gtgcagatga acttcagggt cagcttgccg taggtggcat
     2701 cgccctcacc ctcgccggac acgctgaact tgtggccgtt tacgtcgccg tccagctcga
     2761 ccaggatggg caccaccccg gtgaacagct cctcgccctt gctcaccatg ctccctccgg
     2821 taccgccctt gtacagctcg tccatgccga gagtgatccc ggcggcggtc acgaactcca
     2881 gcaggaccat gtgatcgcgc ttctcgttgg ggtctttcga aagtttggac tgcacgctca
     2941 ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat gggggtgttc tgctggtagt
     3001 ggtaggcgag ctgcacgccg ccgtcctcga tgttgtggcg gatcttgaag ttcgccttga
     3061 tgccgttctt ctgcttgtcg gccttgatat agacgttctc gagtgagctc agccgaccta
     3121 tagctctgac tgcgtgacct gtcttattcc acttacgacg tgatgagtcg accatggtgg
     3181 cgagatcctt atcgtcatcg tcgtacagat cccgacccat ttgctgtcca ccagtcatgc
     3241 tagccatacc atgatgatga tgatgatgag aacccatggt ggcaacagcg ctactagcat
     3301 aacttcgtat aaagtatcct atacgaagtt atttgcctta acccagaaat tatcactgtt
     3361 attctttaga atggtgcaaa gaataacttc gtataatgta tgctatacga agttatgaat
     3421 tccgctcgag ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     3481 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     3541 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt agttcttgcc
     3601 acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc
     3661 actgacaatt ccgtggtgtt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
     3721 atctagcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg
     3781 caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc agggggagat
     3841 gtgggaggtt ttttaaagcg gccgcaggaa cccctagtga tggagttggc cactccctct
     3901 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt
     3961 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg
     4021 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt
     4081 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     4141 ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     4201 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     4261 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
     4321 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     4381 gactcttgtt ccaaactgga acaacactca actctatctc gggctattct tttgatttat
     4441 aagggatttt gccgatttcg gtctattggt taaaaaatga gctgatttaa caaaaattta
     4501 acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct
     4561 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     4621 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     4681 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     4741 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     4801 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     4861 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     4921 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     4981 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     5041 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     5101 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     5161 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     5221 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     5281 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     5341 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     5401 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     5461 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     5521 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     5581 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     5641 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     5701 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     5761 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     5821 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     5881 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     5941 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     6001 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     6061 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     6121 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     6181 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     6241 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     6301 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     6361 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     6421 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     6481 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     6541 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     6601 tgt
//
