LOCUS       sequence-209258-        6907 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector with hSynapsin promoter, Cre-OFF hCAR (for efficient 
            CAV-2 infection), Cre-ON ChETA-mRuby2 (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 6907)
  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 6907)
  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-mRuby2}on-W3SL
FEATURES             Location/Qualifiers
     source          1..6907
                     /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(1939..2649)
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     regulatory      2646..2655
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(2665..3591)
                     /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      3588..3597
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3602..3635
                     /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    3686..3719
                     /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(3789..3809)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4169..4309
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4169..4298
                     /label=AAV2 ITR
     rep_origin      4384..4839
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4471..4490)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4681..4702
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4856..4875)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4975..4997
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5035..5053)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5121..5225
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5226..5294,5295..6086)
                     /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(5444..5463)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6257..6845
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6746..6765
                     /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 gtccatccca ccaccaagcc cggcgaactt
     1981 ggcaactgcg tgttcgcgtt gtactacgaa catttcattg tcactttcct ctaacctttc
     2041 caggcggtga tcaacggcat ggataccggg catcttgatg ttcccgacgg tcttttttga
     2101 cctgtaagtt gttacgaaag agcaagacag atggccacca ccatcaactt tcagtgccat
     2161 atgagtgtat cccctcagac caccatctgc tggatacatc atctctgtat taggctccca
     2221 acccttggtc ttcttctgca tcacgggacc attggaggga aagtttaccc ctctgacttg
     2281 gacgtggtaa acgagacagc catcctcaag gctggtgtcc tgcatgacgg tgacgactcc
     2341 accatcttcg tatctcgtaa ctctttccca agtaaaaccc tcaggaaagg actgtttaaa
     2401 gaaatcagga atgcctttcg ggtacttgat aaaagtacgg ctgccataca tgaacgacgt
     2461 ggcaagaatg tcaaaggcaa atggcagggg tcctccctcg atgactttga tcctcatggt
     2521 ttgagttccc atgtacggat tgccttctcc ttcacctgtg catttgaatt ggtggccgtt
     2581 gaccgaacct tccatgacca ccttcatacg catattttcc ttgatcagct cttcgccctt
     2641 agacaccatg gtggcggccg ctggcacggc tccggcctcg gcttcgtctt cgacgagagt
     2701 ctcgacctcg atctccgttc cgccgatgtt cagtttggtg gttttgcgga tatctccgtg
     2761 aatcaatatg tgctcgtgga tcaggacgcg caggtagtgt cccaacaacc cccaacaatt
     2821 tttactcatc agatcaataa tcgtgtgacc tacggtggag ccatagacgc tcaggacgcc
     2881 aaaaccttcg ggccccaaaa tgaagagaat tgggaacata ccccagctca cgaaaaacag
     2941 ccatgccatg ccggtcacga cctggcggca ccgacccttt ggcacagtat gataaccctc
     3001 gatatatgct ttggcggcgt gaaaaaatgt gttcgcgcca tagcacaatc caagacaaaa
     3061 gaagatgact ttaacatagc cggttgccat ggcgctggta gccccccaca cgatagtccc
     3121 gatgtctgag acaaggagtc ccatggttct cctgctgtag tcgttgctca ggccggtgag
     3181 gttgctcagg cggataagga tgacaggaca agtgagcagc caggttgcat agcgcagcca
     3241 ctgcacccgg tgtcctgtgg caaggtagag catagaggga ttcttaaact caaaaaagaa
     3301 ctcgagaatc accttaacca tttcaatggc gcacacatag atctcctccc agccgcatgt
     3361 agatttccag gtttggtagg catagaacat cagcagcaaa atgctgaatc ctgctgcaag
     3421 ccactgcagg acatttgacg cggtctgagc gccgttcgtg ccgcgagatt caatccatcc
     3481 ggcacagtaa cattgatcct cagggaccag gacggacccg ttcaccacca caggattagt
     3541 aacgaacaaa agttcgcgtc cgacggcaga caaagcgccg ccatagtcca tggtggctag
     3601 cataacttcg tataaagtat cctatacgaa gttatttgcc ttaacccaga aattatcact
     3661 gttattcttt agaatggtgc aaagaataac ttcgtataat gtatgctata cgaagttatg
     3721 aattccgctc gagataatca acctctggat tacaaaattt gtgaaagatt gactggtatt
     3781 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
     3841 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttagttctt
     3901 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     3961 ggcactgaca attccgtggt gtttatttgt gaaatttgtg atgctattgc tttatttgta
     4021 accatctagc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     4081 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga
     4141 gatgtgggag gttttttaaa gcggccgcag gaacccctag tgatggagtt ggccactccc
     4201 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     4261 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg
     4321 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat
     4381 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     4441 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     4501 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     4561 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg
     4621 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     4681 gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt
     4741 tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat
     4801 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa
     4861 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     4921 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     4981 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg
     5041 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg
     5101 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     5161 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     5221 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     5281 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     5341 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5401 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5461 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5521 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5581 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     5641 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     5701 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     5761 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     5821 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     5881 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     5941 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     6001 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     6061 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     6121 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     6181 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     6241 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     6301 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     6361 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     6421 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     6481 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     6541 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     6601 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     6661 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     6721 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     6781 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     6841 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     6901 acatgtt
//
