LOCUS       sequence-238428-        6815 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV plasmid with human synapsin promoter driving mCh and BoNT/B 
            amino acids 1-146 fused to iLID(V416I), separated with an IRES 
            element. Co-express with SSPB-BoNT(147-441, Y365A) for sPA-BoNT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6815)
  AUTHORS   Liu Q, Sinnen BL, Boxer EE, Schneider MW, Grybko MJ, Buchta WC, 
            Gibson ES, Wysoczynski CL, Ford CP, Gottschalk A, Aoto J, Tucker CL,
            Kennedy MJ
  TITLE     A Photoactivatable Botulinum Neurotoxin for Inducible Control of 
            Neurotransmission.
  JOURNAL   Neuron. 2019 Jan 15. pii: S0896-6273(19)30003-0. doi: 
            10.1016/j.neuron.2019.01.002.
  PUBMED    30704911
REFERENCE   2  (bases 1 to 6815)
  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: "Neuron. 
            2019 Jan 15. pii: S0896-6273(19)30003-0. doi: 
            10.1016/j.neuron.2019.01.002."
COMMENT     Sequence Label: pAAV hSyn mCh-IRES-BoNT/B(1-146)-iLID
FEATURES             Location/Qualifiers
     source          1..6815
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   6..135
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        180..627
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      653..662
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             659..1366
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(804..822)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1072..1091
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(1968..1984)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1969..1985)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     CDS             2484..2921
                     /codon_start=1
                     /product="improved light-inducible dimer when used with 
                     SspB nano or SspB micro (Guntas et al., 2015)"
                     /label=iLID
                     /translation="LATTLERIEKNFIITDPRLPDNPIIFASDSFLQLTEYSREEILGR
                     NCRFLQGPETDRATVRKIRDAIDNQTEVTVQLINYTKSGKKFWNVFHLQPMRDYKGDVQ
                     YFIGVQLDGTERLHGAAEREAVCLIKKTAFQIAEAANDENYF"
     misc_feature    2947..3535
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3000..3020)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3418..3429)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3567..4043
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3766..3785)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4083..4223
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4083..4212
                     /label=AAV2 ITR
     rep_origin      4298..4753
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4385..4404)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4595..4616
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4770..4789)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4889..4911
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4949..4967)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5035..5139
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5140..5208,5209..6000)
                     /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(5358..5377)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6171..6759
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6660..6679
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgtcctgc aggcagctgc gcgctcgctc gctcactgag gccgcccggg cgtcgggcga
       61 cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg gccaactcca
      121 tcactagggg ttcctgcggc cgcacgcgtg tgtctagact gcagagggcc ctgcgtatga
      181 gtgcaagtgg gttttaggac caggatgagg cggggtgggg gtgcctacct gacgaccgac
      241 cccgacccac tggacaagca cccaaccccc attccccaaa ttgcgcatcc cctatcagag
      301 agggggaggg gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg
      361 acagtgcctt cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg
      421 ctgacgtcac tcgccggtcc cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc
      481 gccgccgccg gcccagccgg accgcaccac gcgaggcgcg agataggggg gcacgggcgc
      541 gaccatctgc gctgcggcgc cggcgactca gcgctgcctc agtctgcggt gggcagcgga
      601 ggagtcgtgt cgtgcctgag agcgcagtcg agaaggtacc ggatctgcta gcgccaccat
      661 ggtgagcaag ggcgaggagg ataacatggc catcatcaag gagttcatgc gcttcaaggt
      721 gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg
      781 cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc ccctgccctt
      841 cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg tgaagcaccc
      901 cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt
      961 gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc tgcaggacgg
     1021 cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg gccccgtaat
     1081 gcagaagaag accatgggct gggaggcctc ctccgagcgg atgtaccccg aggacggcgc
     1141 cctgaagggc gagatcaagc agaggctgaa gctgaaggac ggcggccact acgacgctga
     1201 ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa
     1261 catcaagttg gacatcacct cccacaacga ggactacacc atcgtggaac agtacgaacg
     1321 cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaagtgtg gcggctagta
     1381 ctccggtatt gcggtaccct tgtacgcctg ttttatactc ccttcccgta acttagacgc
     1441 acaaaaccaa gttcaataga agggggtaca aaccagtacc accacgaaca agcacttctg
     1501 tttccccggt gatgtcgtat agactgcttg cgtggttgaa agcgacggat ccgttatccg
     1561 cttatgtact tcgagaagcc cagtaccacc tcggaatctt cgatgcgttg cgctcagcac
     1621 tcaaccccag agtgtagctt aggctgatga gtctggacat ccctcaccgg tgacggtggt
     1681 ccaggctgcg ttggcggcct acctatggct aacgccatgg gacgctagtt gtgaacaagg
     1741 tgtgaagagc ctattgagct acataagaat cctccggccc ctgaatgcgg ctaatcccaa
     1801 cctcggagca ggtggtcaca aaccagtgat tggcctgtcg taacgcgcaa gtccgtggcg
     1861 gaaccgacta ctttgggtgt ccgtgtttcc ttttatttta ttgtggctgc ttatggtgac
     1921 aatcacagat tgttatcata aagcgaattg gataggatca agcttatcga taccgtcgac
     1981 ctcgagccac catgccagtt acaataaata attttaatta taatgatcct attgataata
     2041 ataatattat tatgatggag cctccatttg cgagaggtac ggggagatat tataaagctt
     2101 ttaaaatcac agatcgtatt tggataatac cggaaagata tacttttgga tataaacctg
     2161 aggattttaa taaaagttcc ggtattttta atagagatgt ttgtgaatat tatgatccag
     2221 attacttaaa tactaatgat aaaaagaata tatttttaca aacaatgatc aagttattta
     2281 atagaatcaa atcaaaacca ttgggtgaaa agttattaga gatgattata aatggtatac
     2341 cttatcttgg agatagacgt gttccactcg aagagtttaa cacaaacatt gctagtgtaa
     2401 ctgttaataa attaatcagt aatccaggag gcggccgcag cggtggcggt ggctctggag
     2461 gtgggtccga gctcggggag tttctggcaa ccacactgga acggatcgag aaaaatttca
     2521 tcattactga tccgagactg cctgacaacc caatcatttt tgcgagcgat tccttcctgc
     2581 agctgacaga atattctcgg gaagagatcc tggggcgcaa ttgccgtttt ctgcagggac
     2641 ccgagacaga ccgtgccact gttcggaaaa tcagagatgc tattgacaac cagactgaag
     2701 tgaccgttca gctgatcaat tataccaaga gcggcaagaa gttctggaac gtgttccacc
     2761 tgcagccgat gcgcgattat aagggcgacg tccagtactt cattggcgtg cagctggatg
     2821 gcaccgaacg tcttcatggc gccgctgagc gtgaggcggt ctgcctgatc aaaaagacag
     2881 cctttcagat tgctgaggca gcgaacgacg aaaattactt ttgaattcga tatcaagctt
     2941 atcgataatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat
     3001 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct
     3061 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag
     3121 gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc
     3181 cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc
     3241 ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     3301 cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg
     3361 ctgctcgcct atgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg
     3421 gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg
     3481 cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcatcga
     3541 taccgagcgc tgctcgagag atctacgggt ggcatccctg tgacccctcc ccagtgcctc
     3601 tcctggccct ggaagttgcc actccagtgc ccaccagcct tgtcctaata aaattaagtt
     3661 gcatcatttt gtctgactag gtgtccttct ataatattat ggggtggagg ggggtggtat
     3721 ggagcaaggg gcaagttggg aagacaacct gtagggcctg cggggtctat tgggaaccaa
     3781 gctggagtgc agtggcacaa tcttggctca ctgcaatctc cgcctcctgg gttcaagcga
     3841 ttctcctgcc tcagcctccc gagttgttgg gattccaggc atgcatgacc aggctcagct
     3901 aatttttgtt tttttggtag agacggggtt tcaccatatt ggccaggctg gtctccaact
     3961 cctaatctca ggtgatctac ccaccttggc ctcccaaatt gctgggatta caggcgtgaa
     4021 ccactgctcc cttccctgtc cttctgattt tgtaggtaac cacgtgcgga ccgagcggcc
     4081 gcaggaaccc ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga
     4141 ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga
     4201 gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg
     4261 cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta
     4321 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccttagcg
     4381 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     4441 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     4501 aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     4561 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     4621 acactcaact ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggtc
     4681 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     4741 acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     4801 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     4861 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     4921 tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     4981 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     5041 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     5101 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     5161 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     5221 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     5281 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     5341 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     5401 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     5461 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     5521 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     5581 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     5641 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     5701 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     5761 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     5821 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     5881 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     5941 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     6001 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     6061 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     6121 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     6181 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     6241 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     6301 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     6361 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     6421 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     6481 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     6541 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     6601 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     6661 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     6721 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     6781 ttacggttcc tggccttttg ctggcctttt gctca
//
