LOCUS       Exported                5791 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV-mediated expression of ultrasensitive protein calcium sensor 
            under the Syn promoter, Cre mediated flip-excision switch.
ACCESSION   .
VERSION     .
KEYWORDS    pGP-AAV-syn-FLEX-jGCaMP7s-WPRE
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5791)
  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 5791)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5791
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    complement(187..220)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    complement(283..316)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     CDS             complement(327..1679)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6s
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRVIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKELFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     primer_bind     886..905
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1145..1166
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1272..1293)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      1574..1583
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     regulatory      1676..1685
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     protein_bind    1696..1729
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    1800..1833
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    1875..2463
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1928..1948)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2346..2357)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2466..2482)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2467..2483)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    2512..2633
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2523..2542)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2577..2596
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3013..3601)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3093..3112)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3772..4632)
                     /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     4395..4414
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4633..4737)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4763..5218)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4900..4921)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5112..5131
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   5290..5419
                     /label=AAV2 ITR
     promoter        5501..157
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
ORIGIN
        1 tcgcgtccgc gccgccgccg gcccagccgg accgcaccac gcgaggcgcg agataggggg
       61 gcacgggcgc gaccatctgc gctgcggcgc cggcgactca gcgctgcctc agtctgcggt
      121 gggcagcgga ggagtcgtgt cgtgcctgag agcgcagtcg aattcaagct gctagcaagg
      181 atccccataa cttcgtataa agtatcctat acgaagttat atcaaaatag gaagaccaat
      241 gcttcaccat cgacccgaat tgccaagcat caccatcgac ccataacttc gtataatgta
      301 tgctatacga agttatacta gagctttcac ttcgctgtca tcatttgtac aaactcttcg
      361 tagtttacct gaccatcccc atcgatgtct gcttccctga tcatttcatc aacctcttca
      421 tctgttaact tctctccaag gtttgtcatc acgtggcgaa gctctgctgc actgatgtag
      481 ccattgccat ccttatcaaa cacaccgaac gcttctctaa tttcttcttc cgtgtccctg
      541 tatttcattt ttcttgccat cattgtcagg aactcaggga agtcgattgt gccgtcaccg
      601 tcggcatcta cttcattgat catgtcctgc agctctgctt ctgtggggtt ctgccccaga
      661 gaccgcatca ccgtccccag ctccttggtt gttattgtcc catccccgtc cttgtcaaat
      721 agggagaaaa gctctttaaa ttctgcgatc tgctcttcag tcagttggtc aggaaggttg
      781 tactccagct tgtgccccag gatgttgccg tcctccttga agtcgatgcc cttaagctcg
      841 atgcggttca ccagggtgtc gccctcgaac ttcacctcgg cgcgggtctt gtagttgccg
      901 tcgtccttga agaagatggt gcgctcctgg atgtagcctt cgggcatggc ggacttgaag
      961 aagtcgtgct gcttcatgtg gtcggggtag cggctgaagc actgcacgcc gtaggtcagg
     1021 gtggtcacga gggtgggcca gggcacgggc agcttgccgg tggtgcagat gaacttcagg
     1081 gtcagcttgc cgtaggtggc atcgccctca ccctcgccgg acacgctgaa cttgtggccg
     1141 tttacgtcgc cgtccagctc gaccaggatg ggcaccaccc cggtgaacag ctcctcgccc
     1201 ttgctcacca tgctccctcc ggtaccgccc ttgtacagct cgtccatgcc gagagtgatc
     1261 ccggcggcgg tcacgaactc cagcaggacc atgtgatcgc gcttctcgtt ggggtctttc
     1321 gaaagtttgg actgcacgct caggtagtgg ttgtcgggca gcagcacggg gccgtcgccg
     1381 atgggggtgt tctgctggta gtggtaggcg agctgcacgc cgccgtcctc gatgttgtgg
     1441 cggatgtgga agttcgcctt gatgccgttc ttctgcttgt cggccttgat atagacgttc
     1501 tcgagtgagc tcagccgacc tatcactctg actgcgtgac ctgtcttatt ccacttacga
     1561 cgtgatgagt cgaccatggt ggcgagatcc ttatcgtcat cgtcgtacag atcccgaccc
     1621 atttgctgtc caccagtcat gctagccata ccatgatgat gatgatgatg agaacccatg
     1681 gtggcggatc ctagtataac ttcgtatagg atactttata cgaagttatc attgggattc
     1741 ttcctatttt gatccaagca tcaccatcga ccctctagtc cagatctcac catcgaccca
     1801 taacttcgta tagcatacat tatacgaagt tatgtccctc gaagaggttc gaattcgata
     1861 tcaagcttat cgataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc
     1921 ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg
     1981 ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc
     2041 tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg
     2101 acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg
     2161 ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga
     2221 caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct
     2281 ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg
     2341 tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc
     2401 ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc
     2461 cgcatcgata ccgtcgacct cgacccgggc ggccgcttcg agcagacatg ataagataca
     2521 ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa
     2581 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca
     2641 acaattgcat tcattttatg tttcaggttc agggggagat gtgggaggtt ttttaaagca
     2701 agtaaaacct ctacaaatgt ggtaaaatcg ataaggatct tcctagagca tggctacgta
     2761 gataagtagc atggcgggtt aatcattaac tacaaggaac ccctagtgat ggagttggcc
     2821 actccctctc tgcgcgctcg ctcgctcact gactncgctg cgctcggtcg ttcggctgcg
     2881 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     2941 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     3001 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     3061 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     3121 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     3181 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     3241 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     3301 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     3361 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     3421 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     3481 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     3541 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     3601 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     3661 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     3721 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     3781 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     3841 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     3901 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     3961 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     4021 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     4081 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     4141 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     4201 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     4261 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     4321 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     4381 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     4441 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     4501 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     4561 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     4621 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     4681 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     4741 atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa attcgcgtta
     4801 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     4861 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     4921 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     4981 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     5041 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     5101 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     5161 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc
     5221 cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta
     5281 ttacgccagc tgcgcgctcg ctcgctcact gaggccgccc gggcaaagcc cgggcgtcgg
     5341 gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg agtggccaac
     5401 tccatcacta ggggttcctt gtagttaatg attaacccgc catgctactt atctacgtag
     5461 ccatgctcta ggaagatctc tgcagagggc cctgcgtatg agtgcaagtg ggttttagga
     5521 ccaggatgag gcggggtggg ggtgcctacc tgacgaccga ccccgaccca ctggacaagc
     5581 acccaacccc cattccccaa attgcgcatc ccctatcaga gagggggagg ggaaacagga
     5641 tgcggcgagg cgcgtgcgca ctgccagctt cagcaccgcg gacagtgcct tcgcccccgc
     5701 ctggcggcgc gcgccaccgc cgcctcagca ctgaaggcgc gctgacgtca ctcgccggtc
     5761 ccccgcaaac tccccttccc ggccaccttg g
//
