LOCUS       sequence-212257-        6901 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector with hSynapsin promoter, Cre-OFF hCAR (for efficient 
            CAV-2 infection), Cre-ON ChETA-EYFP (for optogenetic activation), 
            and W3SL regulatory cassette (for maximize cloning capacity).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6901)
  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 6901)
  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-{ChETA-EYFP}on-W3SL
FEATURES             Location/Qualifiers
     source          1..6901
                     /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="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     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"
     CDS             complement(2660..3586)
                     /codon_start=1
                     /product="H134R mutant of an N-terminal fragment of 
                     channelrhodopsin-2 from the alga Chlamydomonas reinhardtii 
                     (Nagel et al., 2005)"
                     /label=ChR2(H134R)
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYATWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     regulatory      3583..3592
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3597..3630
                     /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    3681..3714
                     /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(3784..3804)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4164..4304
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4164..4293
                     /label=AAV2 ITR
     rep_origin      4379..4834
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4466..4485)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4676..4697
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4851..4870)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4970..4992
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5030..5048)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5116..5220
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5221..5289,5290..6081)
                     /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(5439..5458)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6252..6840
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6741..6760
                     /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 tggtagctca
     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 tagcgggcga agcactgcag gccgtagccg aaggtggtca
     2461 cgagggtggg ccagggcacg ggcagcttgc cggtggtgca gatgaacttc agggtcagct
     2521 tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct gaacttgtgg ccgtttacgt
     2581 cgccgtccag ctcgaccagg atgggcacca ccccggtgaa cagctcctcg cccttgctca
     2641 ccatggtggc ggccgctggc acggctccgg cctcggcttc gtcttcgacg agagtctcga
     2701 cctcgatctc cgttccgccg atgttcagtt tggtggtttt gcggatatct ccgtgaatca
     2761 atatgtgctc gtggatcagg acgcgcaggt agtgtcccaa caacccccaa caatttttac
     2821 tcatcagatc aataatcgtg tgacctacgg tggagccata gacgctcagg acgccaaaac
     2881 cttcgggccc caaaatgaag agaattggga acatacccca gctcacgaaa aacagccatg
     2941 ccatgccggt cacgacctgg cggcaccgac cctttggcac agtatgataa ccctcgatat
     3001 atgctttggc ggcgtgaaaa aatgtgttcg cgccatagca caatccaaga caaaagaaga
     3061 tgactttaac atagccggtt gccatggcgc tggtagcccc ccacacgata gtcccgatgt
     3121 ctgagacaag gagtcccatg gttctcctgc tgtagtcgtt gctcaggccg gtgaggttgc
     3181 tcaggcggat aaggatgaca ggacaagtga gcagccaggt tgcatagcgc agccactgca
     3241 cccggtgtcc tgtggcaagg tagagcatag agggattctt aaactcaaaa aagaactcga
     3301 gaatcacctt aaccatttca atggcgcaca catagatctc ctcccagccg catgtagatt
     3361 tccaggtttg gtaggcatag aacatcagca gcaaaatgct gaatcctgct gcaagccact
     3421 gcaggacatt tgacgcggtc tgagcgccgt tcgtgccgcg agattcaatc catccggcac
     3481 agtaacattg atcctcaggg accaggacgg acccgttcac caccacagga ttagtaacga
     3541 acaaaagttc gcgtccgacg gcagacaaag cgccgccata gtccatggtg gctagcataa
     3601 cttcgtataa agtatcctat acgaagttat ttgccttaac ccagaaatta tcactgttat
     3661 tctttagaat ggtgcaaaga ataacttcgt ataatgtatg ctatacgaag ttatgaattc
     3721 cgctcgagat aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
     3781 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat
     3841 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttag ttcttgccac
     3901 ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac
     3961 tgacaattcc gtggtgttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     4021 ctagctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
     4081 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggagatgt
     4141 gggaggtttt ttaaagcggc cgcaggaacc cctagtgatg gagttggcca ctccctctct
     4201 gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc
     4261 ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc tgatgcggta
     4321 ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac
     4381 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     4441 acacttgcca gcgccttagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     4501 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     4561 gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca
     4621 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     4681 ctcttgttcc aaactggaac aacactcaac tctatctcgg gctattcttt tgatttataa
     4741 gggattttgc cgatttcggt ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     4801 gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc
     4861 tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga
     4921 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     4981 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata
     5041 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     5101 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     5161 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     5221 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     5281 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     5341 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     5401 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     5461 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     5521 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     5581 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     5641 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     5701 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     5761 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     5821 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     5881 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     5941 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     6001 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     6061 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     6121 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     6181 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     6241 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     6301 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     6361 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     6421 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     6481 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     6541 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     6601 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     6661 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     6721 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     6781 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     6841 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     6901 t
//
