LOCUS       sequence-447138-        5624 bp    DNA     circular SYN 14-APR-2025
DEFINITION  AAV expression of ultrasensitive protein calcium sensor from 
            Synapsin promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5624)
  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 5624)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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.GCaMP6s.WPRE.SV40
FEATURES             Location/Qualifiers
     source          1..5624
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   826..955
                     /label=AAV2 ITR
     polyA_signal    1117..1238
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1154..1173)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1208..1227
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1267..1283
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1268..1284
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1287..1875
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1393..1404
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1802..1822
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(1888..2331,2332..2337,2338..2772,2773..2790,
                     2791..3060,3061..3066,3067..3123,3124..3150,3151..3174,
                     3175..3177,3178..3210,3211..3213,3214..3231,3232..3240))
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6s
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     primer_bind     2447..2466
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2706..2727
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2833..2854)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      3135..3144
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      3237..3246
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(3278..3725)
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     repeat_region   3807..3936
                     /label=AAV2 ITR
     rep_origin      4008..4463
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4095..4114)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4305..4326
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4489..4593
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4594..4662,4663..5454)
                     /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(4812..4831)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctgcg cgctcgctcg
      841 ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga cctttggtcg cccggcctca
      901 gtgagcgagc gagcgcgcag agagggagtg gccaactcca tcactagggg ttccttgtag
      961 ttaatgatta acccgccatg ctacttatct acgtagccat gctctaggaa gatccttatc
     1021 gattttacca catttgtaga ggttttactt gctttaaaaa acctcccaca tctccccctg
     1081 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     1141 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     1201 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgctc gaagcggccg
     1261 cccgggtcga ggtcgacggt atcgatgcgg ggaggcggcc caaagggaga tccgactcgt
     1321 ctgagggcga aggcgaagac gcggaagagg ccgcagagcc ggcagcaggc cgcgggaagg
     1381 aaggtccgct ggattgaggg ccgaagggac gtagcagaag gacgtcccgc gcagaatcca
     1441 ggtggcaaca caggcgagca gccaaggaaa ggacgatgat ttccccgaca acaccacgga
     1501 attgtcagtg cccaacagcc gagcccctgt ccagcagcgg gcaaggcagg cggcgatgag
     1561 ttccgccgtg gcaataggga gggggaaagc gaaagtcccg gaaaggagct gacaggtggt
     1621 ggcaatgccc caaccagtgg gggttgcgtc agcaaacaca gtgcacacca cgccacgttg
     1681 cctgacaacg ggccacaact cctcataaag agacagcaac caggatttat acaaggagga
     1741 gaaaatgaaa gccatacggg aagcaatagc atgatacaaa ggcattaaag cagcgtatcc
     1801 acatagcgta aaaggagcaa catagttaag aataccagtc aatctttcac aaattttgta
     1861 atccagaggt tgattatcga taagctttca cttcgctgtc atcatttgta caaactcttc
     1921 gtagtttacc tgaccatccc catcgatgtc tgcttccctg atcatttcat caacctcttc
     1981 atctgttaac ttctctccaa ggtttgtcat cacgtggcga agctctgctg cactgatgta
     2041 gccattgcca tccttatcaa acacaccgaa cgcttctcta atttcttctt ccgtgtccct
     2101 gtatttcatt tttcttgcca tcattgtcag gaactcaggg aagtcgattg tgccgtcacc
     2161 gtcggcatct acttcattga tcatgtcctg cagctctgct tctgtggggt tctgccccag
     2221 agaccgcatc accgtcccca gctccttggt tgttattgtc ccatccccgt ccttgtcaaa
     2281 tagggagaaa gcctctttaa attctgcgat ctgctcttca gtcagttggt ccggcaggtt
     2341 gtactccagc ttgtgcccca ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc
     2401 gatgcggttc accagggtgt cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc
     2461 gtcgtccttg aagaagatgg tgcgctcctg gatgtagcct tcgggcatgg cggacttgaa
     2521 gaagtcgtgc tgcttcatgt ggtcggggta gcggctgaag cactgcacgc cgtaggtcag
     2581 ggtggtcacg agggtgggcc agggcacggg cagcttgccg gtggtgcaga tgaacttcag
     2641 ggtcagcttg ccgtaggtgg catcgccctc accctcgccg gacacgctga acttgtggcc
     2701 gtttacgtcg ccgtccagct cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc
     2761 cttgctcacc atgctccctc cggtaccgcc cttgtacagc tcgtccatgc cgagagtgat
     2821 cccggcggcg gtcacgaact ccagcaggac catgtgatcg cgcttctcgt tggggtcttt
     2881 cgaaagtttg gactgcacgc tcaggtagtg gttgtcgggc agcagcacgg ggccgtcgcc
     2941 gatgggggtg ttctgctggt agtggtaggc gagctgcacg ccgccgtcct cgatgttgtg
     3001 gcggatgtgg aagttcgcct tgatgccgtt cttctgcttg tcggccttga tatagacgtt
     3061 ctcgagtgag ctcagccgac ctatagctct gactgcgtga cctgtcttat tccacttacg
     3121 acgtgatgag tcgaccatgg tggcgagatc cttatcgtca tcgtcgtaca gatcccgacc
     3181 catttgctgt ccaccagtca tgctagccat accatgatga tgatgatgat gagaacccat
     3241 ggtggcgggt ggatccttgc tagcagcttg aattcgactg cgctctcagg cacgacacga
     3301 ctcctccgct gcccaccgca gactgaggca gcgctgagtc gccggcgccg cagcgcagat
     3361 ggtcgcgccc gtgcccccct atctcgcgcc tcgcgtggtg cggtccggct gggccggcgg
     3421 cggcgcggac gcgaccaagg tggccgggaa ggggagtttg cgggggaccg gcgagtgacg
     3481 tcagcgcgcc ttcagtgctg aggcggcggt ggcgcgcgcc gccaggcggg ggcgaaggca
     3541 ctgtccgcgg tgctgaagct ggcagtgcgc acgcgcctcg ccgcatcctg tttcccctcc
     3601 ccctctctga taggggatgc gcaatttggg gaatgggggt tgggtgcttg tccagtgggt
     3661 cggggtcggt cgtcaggtag gcacccccac cccgcctcat cctggtccta aaacccactt
     3721 gcactcatac gcagggccct ctgcagagat cttcctagag catggctacg tagataagta
     3781 gcatggcggg ttaatcatta actacaagga acccctagtg atggagttgg ccactccctc
     3841 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt
     3901 tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgg cgtaatagcg aagaggcccg
     3961 caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg cgccctgtag
     4021 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     4081 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     4141 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     4201 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     4261 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     4321 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     4381 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     4441 caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct
     4501 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     4561 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     4621 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     4681 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     4741 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     4801 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     4861 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     4921 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     4981 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     5041 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     5101 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     5161 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     5221 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     5281 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     5341 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     5401 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     5461 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     5521 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     5581 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttc
//
