LOCUS       sequence-382633-        5491 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses miniSOG targeted to the plasma membrane of neurons, joined
            by a T2A cleavage site with mCherry; for AAV production.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5491)
  AUTHORS   Sessler CD, Zhou Y, Wang W, Hartley ND, Fu Z, Graykowski D, Sheng M,
            Wang X, Liu J
  TITLE     Optogenetic polymerization and assembly of electrically functional 
            polymers for modulation of single-neuron excitability.
  JOURNAL   Sci Adv. 2022 Dec 9;8(49):eade1136. doi: 10.1126/sciadv.ade1136. 
            Epub 2022 Dec 7.
  PUBMED    36475786
REFERENCE   2  (bases 1 to 5491)
  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: "Sci Adv."; 
            date: "2022-12-9"; volume: "8(49):eade1136. doi"; pages: " 
            10.1126/sciadv.ade1136. Epub 2022 Dec 7"
COMMENT     Sequence Label: pAAV-hSyn-mCherry-T2A-miniSOG-C-Far
FEATURES             Location/Qualifiers
     source          1..5491
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        26..473
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     CDS             504..1211
                     /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(649..667)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     917..936
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1227..1280
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             1290..1313
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1329..1646
                     /codon_start=1
                     /product="mini singlet oxygen generator, a fluorescent 
                     flavoprotein suitable for correlated light and electron 
                     microscopy (Shu et al., 2011)"
                     /label=miniSOG
                     /translation="MEKSFVITDPRLPDNPIIFASDGFLELTEYSREEILGRNGRFLQG
                     PETDQATVQKIRDAIRDQREITVQLINYTKSGKKFWNLLHLQPMRDQKGELQYFIGVQL
                     DG"
     misc_feature    1743..2331
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1796..1816)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2214..2225)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2334..2350)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2335..2351)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    2380..2501
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2391..2410)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2445..2464
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2663..2792
                     /label=AAV2 ITR
     rep_origin      2864..3319
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2951..2970)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3161..3182
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3345..3449
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3450..3518,3519..4310)
                     /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(3668..3687)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4481..5069
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4970..4989
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   5306..5435
                     /label=AAV2 ITR
ORIGIN
        1 atctctgcag agggccctgc gtatgagtgc aagtgggttt taggaccagg atgaggcggg
       61 gtgggggtgc ctacctgacg accgaccccg acccactgga caagcaccca acccccattc
      121 cccaaattgc gcatccccta tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt
      181 gcgcactgcc agcttcagca ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc
      241 accgccgcct cagcactgaa ggcgcgctga cgtcactcgc cggtcccccg caaactcccc
      301 ttcccggcca ccttggtcgc gtccgcgccg ccgccggccc agccggaccg caccacgcga
      361 ggcgcgagat aggggggcac gggcgcgacc atctgcgctg cggcgccggc gactcagcgc
      421 tgcctcagtc tgcggtgggc agcggaggag tcgtgtcgtg cctgagagcg cagtcgaatt
      481 caagctgcta gcaaggatcc ggtatggtga gcaagggcga ggaggataac atggccatca
      541 tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg
      601 agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg
      661 tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg
      721 gctccaaggc ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc
      781 ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga
      841 cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca
      901 acttcccctc cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg
      961 agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga
     1021 aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc
     1081 agctgcccgg cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact
     1141 acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg
     1201 agctgtacaa gggatctggt ggcggtgagg gcagaggaag tcttctaaca tgcggtgacg
     1261 tggaggagaa tcccggccct cgtacgatgg attacaagga tgacgacgat aagggcggag
     1321 gtggttctat ggagaaaagt ttcgtgataa ctgatccacg gctgccagac aatcccatca
     1381 tcttcgcatc cgatggcttc ctggagctga ccgagtattc cagagaggag atcctgggcc
     1441 gcaatggccg ctttctgcag ggaccagaga cagaccaggc cacagtgcag aagattcgcg
     1501 atgccattag agatcagcgc gagattaccg tgcagctgat aaactacaca aaaagcggga
     1561 agaaattctg gaacctcctg cacctccagc ccatgaggga ccagaagggt gagctccagt
     1621 atttcatcgg agtgcagctg gatggatcag gaggaggctc aggaggagga aagctgaacc
     1681 ctcctgatga gagtggcccc ggctgcatga gctgctgtgt gctctcctaa aagcttatcg
     1741 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     1801 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     1861 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
     1921 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
     1981 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
     2041 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
     2101 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
     2161 tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
     2221 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
     2281 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc
     2341 gtcgacctcg acccgggcgg ccgcttcgag cagacatgat aagatacatt gatgagtttg
     2401 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     2461 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     2521 attttatgtt tcaggttcag ggggagatgt gggaggtttt ttaaagcaag taaaacctct
     2581 acaaatgtgg taaaatcgat aaggatcttc ctagagcatg gctacgtaga taagtagcat
     2641 ggcgggttaa tcattaacta caaggaaccc ctagtgatgg agttggccac tccctctctg
     2701 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     2761 cgggcggcct cagtgagcga gcgagcgcgc agctggcgta atagcgaaga ggcccgcacc
     2821 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc
     2881 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     2941 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     3001 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     3061 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     3121 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     3181 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     3241 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     3301 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     3361 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     3421 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     3481 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     3541 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     3601 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     3661 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     3721 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     3781 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     3841 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     3901 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     3961 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     4021 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     4081 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     4141 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     4201 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     4261 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     4321 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     4381 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     4441 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     4501 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4561 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     4621 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     4681 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     4741 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     4801 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     4861 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     4921 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     4981 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     5041 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     5101 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     5161 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     5221 gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg
     5281 cgcgttggcc gattcattaa tgcagctgcg cgctcgctcg ctcactgagg ccgcccgggc
     5341 aaagcccggg cgtcgggcga cctttggtcg cccggcctca gtgagcgagc gagcgcgcag
     5401 agagggagtg gccaactcca tcactagggg ttccttgtag ttaatgatta acccgccatg
     5461 ctacttatct acgtagccat gctctaggaa g
//
