LOCUS       sequence-375715-        6243 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector for Cre- and Flp-dependent transgene expression (CRE-ON /
            FLP-OFF) of ChR2-YFP in neurons under the control of the hSyn1 
            promoter (Kugler et al 2003).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6243)
  AUTHORS   Pouchelon G, Vergara J, McMahon J, Gorissen BL, Lin JD, 
            Vormstein-Schneider D, Niehaus JL, Burbridge TJ, Wester JC, Sherer 
            M, Fernandez-Otero M, Allaway KC, Pelkey K, Chittajallu R, McBain 
            CJ, Fan M, Nasse JS, Wildenberg GA, Fishell G, Dimidschstein J
  TITLE     A versatile viral toolkit for functional discovery in the nervous 
            system.
  JOURNAL   Cell Rep Methods. 2022 May 26;2(6):100225. doi: 
            10.1016/j.crmeth.2022.100225. eCollection 2022 Jun 20.
  PUBMED    35784651
REFERENCE   2  (bases 1 to 6243)
  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"; journalName: "Cell Rep 
            Methods."; date: "2022-05-26"; volume: "2(6):100225. doi"; pages: " 
            10.1016/j.crmeth.2022.100225. eCollection 2022 Jun 20"
COMMENT     Sequence Label: pAAV-VTKS5-ChR2-YFP
FEATURES             Location/Qualifiers
     source          1..6243
                     /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        158..605
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    629..662
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     protein_bind    complement(669..702)
                     /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(753..786)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(807..1520,1521..1523,1524..1526))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(852..873)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1201..1220
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1460..1481
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1523..1532
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(1542..2468)
                     /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
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     regulatory      2465..2474
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    2524..2557
                     /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    2608..2641
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2642..2675
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    2684..3272
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2737..2757)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3155..3166)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3280..3504
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3506..3646
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3506..3635
                     /label=AAV2 ITR
     rep_origin      3721..4176
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3808..3827)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4018..4039
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4193..4212)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4312..4334
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4372..4390)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4458..4562
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4563..4631,4632..5423)
                     /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(4781..4800)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5594..6182
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6083..6102
                     /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 gctagactgc agagggccct gcgtatgagt gcaagtgggt tttaggacca
      181 ggatgaggcg gggtgggggt gcctacctga cgaccgaccc cgacccactg gacaagcacc
      241 caacccccat tccccaaatt gcgcatcccc tatcagagag ggggagggga aacaggatgc
      301 ggcgaggcgc gtgcgcactg ccagcttcag caccgcggac agtgccttcg cccccgcctg
      361 gcggcgcgcg ccaccgccgc ctcagcactg aaggcgcgct gacgtcactc gccggtcccc
      421 cgcaaactcc ccttcccggc caccttggtc gcgtccgcgc cgccgccggc ccagccggac
      481 cgcaccacgc gaggcgcgag ataggggggc acgggcgcga ccatctgcgc tgcggcgccg
      541 gcgactcagc gctgcctcag tctgcggtgg gcagcggagg agtcgtgtcg tgcctgagag
      601 cgcagtcaga tatcgtttaa actccggaga agttcctatt ctctagaaag tataggaact
      661 tctccggaat aacttcgtat aggatacttt atacgaagtt atgcagaatg gtagctggat
      721 tgtagctgct attagcaata tgaaacctct taataacttc gtatagcata cattatacga
      781 agttataacg cgccactagt gaattcttac ttgtacagct cgtccatgcc gagagtgatc
      841 ccggcggcgg tcacgaactc cagcaggacc atgtgatcgc gcttctcgtt ggggtctttg
      901 ctcagggcgg actggtagct caggtagtgg ttgtcgggca gcagcacggg gccgtcgccg
      961 atgggggtgt tctgctggta gtggtcggcg agctgcacgc tgccgtcctc gatgttgtgg
     1021 cggatcttga agttcacctt gatgccgttc ttctgcttgt cggccatgat atagacgttg
     1081 tggctgttgt agttgtactc cagcttgtgc cccaggatgt tgccgtcctc cttgaagtcg
     1141 atgcccttca gctcgatgcg gttcaccagg gtgtcgccct cgaacttcac ctcggcgcgg
     1201 gtcttgtagt tgccgtcgtc cttgaagaag atggtgcgct cctggacgta gccttcgggc
     1261 atggcggact tgaagaagtc gtgctgcttc atgtggtcgg ggtagcgggc gaagcactgc
     1321 aggccgtagc cgaaggtggt cacgagggtg ggccagggca cgggcagctt gccggtggtg
     1381 cagatgaact tcagggtcag cttgccgtag gtggcatcgc cctcgccctc gccggacacg
     1441 ctgaacttgt ggccgtttac gtcgccgtcc agctcgacca ggatgggcac caccccggtg
     1501 aacagctcct cgcccttgct caccatggtg gcggccgctg gcacggctcc ggcctcggct
     1561 tcgtcttcga cgagagtctc gacctcgatc tccgttccgc cgatgttcag tttggtggtt
     1621 ttgcggatat ctccgtgaat caatatgtgc tcgtggatca ggacgcgcag gtagtgtccc
     1681 aacaaccccc aacaattttt actcatcaga tcaataatcg tgtgacctac ggtggagcca
     1741 tagacgctca ggacgccaaa accttcgggc cccaaaatga agagaattgg gaacataccc
     1801 cagctcacga aaaacagcca tgccatgccg gtcacgacct ggcggcaccg accctttggc
     1861 acagtatgat aaccctcgat atatgctttg gcggcgtgaa aaaatgtgtt cgcgccatag
     1921 cacaatccaa gacaaaagaa gatgacttta acatagccgg ttgccatggc gctggtagcc
     1981 ccccacacga tagtcccgat gtctgagaca aggagtccca tggttctcct gctgtagtcg
     2041 ttgctcaggc cggtgaggtt gctcaggcgg ataaggatga caggacaagt gagcagccac
     2101 tctgcatagc gcagccactg cacccggtgt cctgtggcaa ggtagagcat agagggattc
     2161 ttaaactcaa aaaagaactc gagaatcacc ttaaccattt caatggcgca cacatagatc
     2221 tcctcccagc cgcatgtaga tttccaggtt tggtaggcat agaacatcag cagcaaaatg
     2281 ctgaatcctg ctgcaagcca ctgcaggaca tttgacgcgg tctgagcgcc gttcgtgccg
     2341 cgagattcaa tccatccggc acagtaacat tgatcctcag ggaccaggac ggacccgttc
     2401 accaccacag gattagtaac gaacaaaagt tcgcgtccga cggcagacaa agcgccgcca
     2461 tagtccatgg tggctccgga gtcgacctgc aggcatgcaa gctttactag tggtggcgct
     2521 agcataactt cgtataaagt atcctatacg aagttatttg ccttaaccca gaaattatca
     2581 ctgttattct ttagaatggt gcaaagaata acttcgtata atgtatgcta tacgaagtta
     2641 tgaagttcct attctctaga aagtatagga acttcggcgc gccaatcaac ctctggatta
     2701 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     2761 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     2821 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     2881 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     2941 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     3001 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     3061 cgtggtgttg tcggggaagc tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg
     3121 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     3181 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac
     3241 gagtcggatc tccctttggg ccgcctcccc gcctggatcc tgtgccttct agttgccagc
     3301 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     3361 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     3421 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     3481 ctggggatgc ggtgggctct atggcaggaa cccctagtga tggagttggc cactccctct
     3541 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt
     3601 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg
     3661 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt
     3721 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     3781 ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     3841 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     3901 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
     3961 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     4021 gactcttgtt ccaaactgga acaacactca actctatctc gggctattct tttgatttat
     4081 aagggatttt gccgatttcg gtctattggt taaaaaatga gctgatttaa caaaaattta
     4141 acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct
     4201 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     4261 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     4321 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     4381 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     4441 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     4501 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     4561 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     4621 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     4681 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     4741 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     4801 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     4861 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     4921 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     4981 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     5041 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     5101 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     5161 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     5221 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     5281 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     5341 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     5401 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     5461 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     5521 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     5581 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     5641 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     5701 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     5761 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     5821 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     5881 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     5941 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     6001 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     6061 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     6121 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     6181 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     6241 tgt
//
