LOCUS       sequence-198300-        5917 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV expression of Cre recombinase-activated ultrasensitive protein 
            calcium sensor from Synapsin promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5917)
  AUTHORS   Chen TW, Wardill TJ, Sun Y, Pulver SR, Renninger SL, Baohan A, 
            Schreiter ER, Kerr RA, Orger MB, Jayaraman V, Looger LL, Svoboda K, 
            Kim DS
  TITLE     Ultrasensitive fluorescent proteins for imaging neuronal activity.
  JOURNAL   Nature. 2013 Jul 18;499(7458):295-300. doi: 10.1038/nature12354.
  PUBMED    23868258
REFERENCE   2  (bases 1 to 5917)
  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"; journalName: 
            "Nature"; date: "2013-07-18- 18"; volume: "499"; issue: "7458"; 
            pages: "295-300"
COMMENT     Sequence Label: pAAV.Syn.Flex.GCaMP6m.WPRE.SV40
FEATURES             Location/Qualifiers
     source          1..5917
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        23..470
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    500..533
                     /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    596..629
                     /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(640..1083,1084..1089,1090..1524,1525..1542,
                     1543..1812,1813..1818,1819..1875,1876..1902,1903..1926,
                     1927..1929,1930..1962,1963..1965,1966..1983,1984..1992))
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6m
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKGSYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     primer_bind     1199..1218
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1458..1479
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1585..1606)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      1887..1896
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1989..1998
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(2009..2042)
                     /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(2113..2146)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2188..2776
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2241..2261)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2659..2670)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2779..2795)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2780..2796)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    2825..2946
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2836..2855)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2890..2909
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3452..4040)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3532..3551)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4211..5002,5003..5071))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRXDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPXAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4834..4853
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5072..5176)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5202..5657)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5339..5360)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5551..5570
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   5729..5858
                     /label=AAV2 ITR
ORIGIN
        1 tctgcagagg gccctgcgta tgagtgcaag tgggttttag gaccaggatg aggcggggtg
       61 ggggtgccta cctgacgacc gaccccgacc cactggacaa gcacccaacc cccattcccc
      121 aaattgcgca tcccctatca gagaggggga ggggaaacag gatgcggcga ggcgcgtgcg
      181 cactgccagc ttcagcaccg cggacagtgc cttcgccccc gcctggcggc gcgcgccacc
      241 gccgcctcag cactgaaggc gcgctgacgt cactcgccgg tcccccgcaa actccccttc
      301 ccggccacct tggtcgcgtc cgcgccgccg ccggcccagc cggaccgcac cacgcgaggc
      361 gcgagatagg ggggcacggg cgcgaccatc tgcgctgcgg cgccggcgac tcagcgctgc
      421 ctcagtctgc ggtgggcagc ggaggagtcg tgtcgtgcct gagagcgcag tcgaattcaa
      481 gctgctagca aggatcccca taacttcgta taaagtatcc tatacgaagt tatatcaaaa
      541 taggaagacc aatgcttcac catcgacccg aattgccaag catcaccatc gacccataac
      601 ttcgtataat gtatgctata cgaagttata ctagagcttt cacttcgctg tcatcatttg
      661 tacaaactct tcgtagttta cctgaccatc cccatcgatg tctgcttccc tgatcatttc
      721 atcaacctct tcatctgtta acttctctcc aaggtttgtc atcacgtggc gaagctctgc
      781 tgcactgatg tagccattgc catccttatc aaacacaccg aacgcttctc taatttcttc
      841 ttccgtgtcc ctgtagctcc cttttcttgc catcattgtc aggaactcag ggaagtcgat
      901 tgtgccgtca ccgtcggcat ctacttcatt gatcatgtcc tgcagctctg cttctgtggg
      961 gttctgcccc agagaccgca tcaccgtccc cagctccttg gttgttattg tcccatcccc
     1021 gtccttgtca aatagggaga aagcctcttt aaattctgcg atctgctctt cagtcagttg
     1081 gtccggcagg ttgtactcca gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat
     1141 gcccttcagc tcgatgcggt tcaccagggt gtcgccctcg aacttcacct cggcgcgggt
     1201 cttgtagttg ccgtcgtcct tgaagaagat ggtgcgctcc tggatgtagc cttcgggcat
     1261 ggcggacttg aagaagtcgt gctgcttcat gtggtcgggg tagcggctga agcactgcac
     1321 gccgtaggtc agggtggtca cgagggtggg ccagggcacg ggcagcttgc cggtggtgca
     1381 gatgaacttc agggtcagct tgccgtaggt ggcatcgccc tcaccctcgc cggacacgct
     1441 gaacttgtgg ccgtttacgt cgccgtccag ctcgaccagg atgggcacca ccccggtgaa
     1501 cagctcctcg cccttgctca ccatgctccc tccggtaccg cccttgtaca gctcgtccat
     1561 gccgagagtg atcccggcgg cggtcacgaa ctccagcagg accatgtgat cgcgcttctc
     1621 gttggggtct ttcgaaagtt tggactgcac gctcaggtag tggttgtcgg gcagcagcac
     1681 ggggccgtcg ccgatggggg tgttctgctg gtagtggtag gcgagctgca cgccgccgtc
     1741 ctcgatgttg tggcggatct tgaagttcgc cttgatgccg ttcttctgct tgtcggcctt
     1801 gatatagacg ttctcgagtg agctcagccg acctatagct ctgactgcgt gacctgtctt
     1861 attccactta cgacgtgatg agtcgaccat ggtggcgaga tccttatcgt catcgtcgta
     1921 cagatcccga cccatttgct gtccaccagt catgctagcc ataccatgat gatgatgatg
     1981 atgagaaccc atggtggcgg atcctagtat aacttcgtat aggatacttt atacgaagtt
     2041 atcattggga ttcttcctat tttgatccaa gcatcaccat cgaccctcta gtccagatct
     2101 caccatcgac ccataacttc gtatagcata cattatacga agttatgtcc ctcgaagagg
     2161 ttcgaattcg atatcaagct tatcgataat caacctctgg attacaaaat ttgtgaaaga
     2221 ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
     2281 cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
     2341 tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
     2401 actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
     2461 tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
     2521 gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
     2581 aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
     2641 tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
     2701 ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
     2761 tgggccgcct ccccgcatcg ataccgtcga cctcgacccg ggcggccgct tcgagcagac
     2821 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
     2881 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     2941 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag
     3001 gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga tcttcctaga
     3061 gcatggctac gtagataagt agcatggcgg gttaatcatt aactacaagg aacccctagt
     3121 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg cccgggcttt
     3181 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgca ttaatgaatc ggccaacgcg
     3241 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     3301 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     3361 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     3421 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     3481 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     3541 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     3601 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     3661 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     3721 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     3781 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     3841 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agracagtat
     3901 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     3961 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     4021 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     4081 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     4141 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     4201 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     4261 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     4321 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     4381 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     4441 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     4501 gtttgcgcaa cgttgttgcc attgctrcag gcatcgtggt gtcacgctcg tcgtttggta
     4561 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     4621 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     4681 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     4741 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     4801 gaccgagttg ctcttgcccg gcgtcaayac gggataatac cgcgccacat agcagaactt
     4861 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     4921 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     4981 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     5041 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     5101 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     5161 aaataggggt tccgcgcaca tttccccgaa aagtgccacc taaattgtaa gcgttaatat
     5221 tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
     5281 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc
     5341 agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac
     5401 cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc
     5461 gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg
     5521 gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag
     5581 ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc
     5641 gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg aagggcgatc
     5701 ggtgcgggcc tcttcgctat tacgccagct gcgcgctcgc tcgctcactg aggccgcccg
     5761 ggcaaagccc gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg
     5821 cagagaggga gtggccaact ccatcactag gggttccttg tagttaatga ttaacccgcc
     5881 atgctactta tctacgtagc catgctctag gaagatc
//
