LOCUS       sequence-200290-        5788 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of ultrasensitive protein calcium sensor 
            under the Syn promoter, Cre mediated flip-excision switch.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5788)
  AUTHORS   Dana H, Sun Y, Mohar B, Hulse BK, Kerlin AM, Hasseman JP, Tsegaye G,
            Tsang A, Wong A, Patel R, Macklin JJ, Chen Y, Konnerth A, Jayaraman 
            V, Looger LL, Schreiter ER, Svoboda K, Kim DS
  TITLE     High-performance calcium sensors for imaging activity in neuronal 
            populations and microcompartments.
  JOURNAL   Nat Methods. 2019 Jul;16(7):649-657. doi: 10.1038/s41592-019-0435-6.
            Epub 2019 Jun 17.
  PUBMED    31209382
REFERENCE   2  (bases 1 to 5788)
  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: 
            "10.1038/s41592-019-0435-6"; journalName: "Nat Methods"; date: 
            "2019-07"; volume: "16"; issue: "7"; pages: "649-657"
COMMENT     Sequence Label: pGP-AAV-syn-FLEX-jGCaMP7b-WPRE
FEATURES             Location/Qualifiers
     source          1..5788
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   45..174
                     /label=AAV2 ITR
     promoter        256..703
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    733..766
                     /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    829..862
                     /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(873..1316,1317..1322,1323..1757,1758..1775,
                     1776..2045,2046..2051,2052..2108,2109..2135,2136..2159,
                     2160..2162,2163..2195,2196..2198,2199..2216,2217..2225))
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNPPDQLTEEQIAEFKELFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMYARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     primer_bind     1432..1451
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1691..1712
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1818..1839)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      2120..2129
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2222..2231
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(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    complement(2346..2379)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2421..3009
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2474..2494)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2892..2903)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3012..3028)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3013..3029)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    3058..3179
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3069..3088)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3123..3142
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3556..4144)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3636..3655)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4315..5106,5107..5175))
                     /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     4938..4957
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5176..5280)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5306..5761)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5443..5464)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5655..5674
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctgcgc gctcgctcgc
       61 tcactgaggc cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag
      121 tgagcgagcg agcgcgcaga gagggagtgg ccaactccat cactaggggt tccttgtagt
      181 taatgattaa cccgccatgc tacttatcta cgtagccatg ctctaggaag atctctgcag
      241 agggccctgc gtatgagtgc aagtgggttt taggaccagg atgaggcggg gtgggggtgc
      301 ctacctgacg accgaccccg acccactgga caagcaccca acccccattc cccaaattgc
      361 gcatccccta tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt gcgcactgcc
      421 agcttcagca ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc accgccgcct
      481 cagcactgaa ggcgcgctga cgtcactcgc cggtcccccg caaactcccc ttcccggcca
      541 ccttggtcgc gtccgcgccg ccgccggccc agccggaccg caccacgcga ggcgcgagat
      601 aggggggcac gggcgcgacc atctgcgctg cggcgccggc gactcagcgc tgcctcagtc
      661 tgcggtgggc agcggaggag tcgtgtcgtg cctgagagcg cagtcgaatt caagctgcta
      721 gcaaggatcc ccataacttc gtataaagta tcctatacga agttatatca aaataggaag
      781 accaatgctt caccatcgac ccgaattgcc aagcatcacc atcgacccat aacttcgtat
      841 aatgtatgct atacgaagtt atactagagc tttcacttcg ctgtcatcat ttgtacaaac
      901 tcttcgtagt ttacctgacc atccccatcg atgtctgctt ccctgatcat ttcatcaacc
      961 tcttcatctg ttaacttctc tccaaggttt gtcatcacgt ggcgaagctc tgctgcactg
     1021 atgtagccat tgccatcctt atcaaacaca ccgaacgctt ctctaatttc ttcttccgtg
     1081 tccctgtatt tcatttttct tgcgtacatt gtcaggaact cagggaagtc gattgtgccg
     1141 tcaccgtcgg catctacttc attgatcatg tcctgcagct ctgcttctgt ggggttctgc
     1201 cccagagacc gcatcaccgt ccccagctcc ttggttgtta ttgtcccatc cccgtccttg
     1261 tcaaataggg agaaaagctc tttaaattct gcgatctgct cttcagtcag ttggtcaggt
     1321 gggttgtact ccagcttgtg ccccaggatg ttgccgtcct ccttgaagtc gatgccctta
     1381 agctcgatgc ggttcaccag ggtgtcgccc tcgaacttca cctcggcgcg ggtcttgtag
     1441 ttgccgtcgt ccttgaagaa gatggtgcgc tcctggatgt agccttcggg catggcggac
     1501 ttgaagaagt cgtgctgctt catgtggtcg gggtagcggc tgaagcactg cacgccgtag
     1561 gtcagggtgg tcacgagggt gggccagggc acgggcagct tgccggtggt gcagatgaac
     1621 ttcagggtca gcttgccgta ggtggcatcg ccctcaccct cgccggacac gctgaacttg
     1681 tggccgttta cgtcgccgtc cagctcgacc aggatgggca ccaccccggt gaacagctcc
     1741 tcgcccttgc tcaccatgct ccctccggta ccgcccttgt acagctcgtc catgccgaga
     1801 gtgatcccgg cggcggtcac gaactccagc aggaccatgt gatcgcgctt ctcgttgggg
     1861 tctttcgaaa gtttggactg cacgctcagg tagtggttgt cgggcagcag cacggggccg
     1921 tcgccgatgg gggtgttctg ctggtagtgg taggcgagct gcacgccgcc gtcctcgatg
     1981 ttgtggcgga tcttgaagtt cgccttgatg ccgttcttct gcttgtcggc cttgatatag
     2041 acgttctcga gtgagctcag ccgacctata gctctgactg cgtgacctgt cttattccac
     2101 ttacgacgtg atgagtcgac catggtggcg agatccttat cgtcatcgtc gtacagatcc
     2161 cgacccattt gctgtccacc agtcatgcta gccataccat gatgatgatg atgatgagaa
     2221 cccatggtgg cggatcctag tataacttcg tataggatac tttatacgaa gttatcattg
     2281 ggattcttcc tattttgatc caagcatcac catcgaccct ctagtccaga tctcaccatc
     2341 gacccataac ttcgtatagc atacattata cgaagttatg tccctcgaag aggttcgaat
     2401 tcgatatcaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg
     2461 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     2521 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
     2581 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
     2641 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
     2701 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
     2761 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat
     2821 cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct
     2881 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     2941 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     3001 cctccccgca tcgataccgt cgacctcgac ccgggcggcc gcttcgagca gacatgataa
     3061 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
     3121 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
     3181 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggagatgtgg gaggtttttt
     3241 aaagcaagta aaacctctac aaatgtggta aaatcgataa ggatcttcct agagcatggc
     3301 tacgtagata agtagcatgg cgggttaatc attaactaca aggaacccct agtgatggag
     3361 ttggccactc cctctctgcg cgctcgctcr ctsactcgct gcgctcggtc gttcggctgn
     3421 cggcgagcgg tatcagctca ctncaaaggc ggtaatacgg ttatccacag aatcagggga
     3481 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     3541 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     3601 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     3661 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     3721 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     3781 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     3841 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     3901 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     3961 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     4021 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     4081 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     4141 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     4201 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     4261 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     4321 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     4381 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     4441 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     4501 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     4561 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     4621 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     4681 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     4741 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     4801 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     4861 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     4921 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     4981 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     5041 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     5101 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     5161 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     5221 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     5281 cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg
     5341 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     5401 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     5461 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     5521 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     5581 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     5641 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     5701 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     5761 tcccattcgc cattcaggct gcgcaact
//
