LOCUS       sequence-355042-        5961 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector for Cre- and Flp-dependent transgene expression (CRE-ON /
            FLP-OFF) of GCaMP6f 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 5961)
  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 5961)
  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-GCaMP6f
FEATURES             Location/Qualifiers
     source          1..5961
                     /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(840..1283,1284..1289,1290..1724,1725..1742,
                     1743..2012,2013..2018,2019..2075,2076..2102,2103..2126,
                     2127..2129,2130..2162,2163..2165,2166..2183,2184..2192))
                     /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     1399..1418
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1658..1679
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1785..1806)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      2087..2096
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2189..2198
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    2242..2275
                     /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    2326..2359
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2360..2393
                     /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    2402..2990
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2455..2475)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2873..2884)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2998..3222
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3224..3364
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3224..3353
                     /label=AAV2 ITR
     rep_origin      3439..3894
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3526..3545)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3736..3757
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3911..3930)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4030..4052
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4090..4108)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4176..4280
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4281..4349,4350..5141)
                     /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(4499..4518)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5312..5900
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5801..5820
                     /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 gaattcgagc tcggtacccg gggatcctct agagtcgact
      841 cacttcgctg tcatcatttg tacaaactct tcgtagttta cctgaccatc cccatcgatg
      901 tctgcttccc tgatcatttc atcaacctct tcatctgtta acttctctcc aaggtttgtc
      961 atcacgtggc gaagctctgc tgcactgatg tagccattgc catccttatc aaacacaccg
     1021 aacgcttctc taatttcttc ttccgtgtcc ctgtatttca tttttcttgc catcattgtc
     1081 aggaactcag ggaagtcgat tgtgccgtca ccgtcggcat ctacttcatt gatcatgtcc
     1141 tgcagctctg cttctgtggg gttctgcccc agagaccgca tcaccgtccc cagctccttg
     1201 gttgttattg tcccatcccc gtccttgtca aatagggaga attcctcttt aaattctgcg
     1261 atctgctctt cagtcagttg gtccggcagg ttgtactcca gcttgtgccc caggatgttg
     1321 ccgtcctcct tgaagtcgat gcccttcagc tcgatgcggt tcaccagggt gtcgccctcg
     1381 aacttcacct cggcgcgggt cttgtagttg ccgtcgtcct tgaagaagat ggtgcgctcc
     1441 tggatgtagc cttcgggcat ggcggacttg aagaagtcgt gctgcttcat gtggtcgggg
     1501 tagcggctga agcactgcac gccgtaggtc agggtggtca cgagggtggg ccagggcacg
     1561 ggcagcttgc cggtggtgca gatgaacttc agggtcagct tgccgtaggt ggcatcgccc
     1621 tcaccctcgc cggacacgct gaacttgtgg ccgtttacgt cgccgtccag ctcgaccagg
     1681 atgggcacca ccccggtgaa cagctcctcg cccttgctca ccatgctccc tccggtaccg
     1741 cccttgtaca gctcgtccat gccgagagtg atcccggcgg cggtcacgaa ctccagcagg
     1801 accatgtgat cgcgcttctc gttggggtct ttcgaaagtt tggactgcac gctcaggtag
     1861 tggttgtcgg gcagcagcac ggggccgtcg ccgatggggg tgttctgctg gtagtggtag
     1921 gcgagctgca cgccgccgtc ctcgatgttg tggcggatct tgaagttcgc cttgatgccg
     1981 ttcttctgct tgtcggcctt gatatagacg ttctcgagtg agctcagccg acctatagct
     2041 ctgactgcgt gacctgtctt attccactta cgacgtgatg agtcgaccat ggtggcgaga
     2101 tccttatcgt catcgtcgta cagatcccga cccatttgct gtccaccagt catgctagcc
     2161 ataccatgat gatgatgatg atgagaaccc atggtggcgt cgacctgcag gcatgcaagc
     2221 tttactagtg gtggcgctag cataacttcg tataaagtat cctatacgaa gttatttgcc
     2281 ttaacccaga aattatcact gttattcttt agaatggtgc aaagaataac ttcgtataat
     2341 gtatgctata cgaagttatg aagttcctat tctctagaaa gtataggaac ttcggcgcgc
     2401 caatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc
     2461 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg
     2521 tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt
     2581 gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac
     2641 tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc
     2701 tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct
     2761 gttgggcact gacaattccg tggtgttgtc ggggaagctg acgtcctttc catggctgct
     2821 cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct
     2881 caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct
     2941 tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc ctggatcctg
     3001 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     3061 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     3121 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     3181 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcaggaacc cctagtgatg
     3241 gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc
     3301 gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg
     3361 caggggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     3421 cgtcaaagca accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     3481 ttacgcgcag cgtgaccgct acacttgcca gcgccttagc gcccgctcct ttcgctttct
     3541 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     3601 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg
     3661 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     3721 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac tctatctcgg
     3781 gctattcttt tgatttataa gggattttgc cgatttcggt ctattggtta aaaaatgagc
     3841 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt
     3901 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa
     3961 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     4021 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     4081 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     4141 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     4201 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     4261 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     4321 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     4381 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     4441 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     4501 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     4561 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     4621 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     4681 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     4741 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     4801 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     4861 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     4921 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     4981 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     5041 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     5101 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     5161 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     5221 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     5281 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     5341 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5401 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     5461 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     5521 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     5581 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     5641 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     5701 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     5761 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     5821 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     5881 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5941 gctggccttt tgctcacatg t
//
