LOCUS       sequence-410393-        7237 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of the inhibitory chemogenetic tool PSAM4-GlyR and 
            mCherry reporter. Application of ultrapotent agonists induces 
            neuronal silencing..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7237)
  AUTHORS   Perez-Sanchez J, Middleton SJ, Pattison LA, Hilton H, Ali 
            Awadelkareem M, Zuberi SR, Renke MB, Hu H, Yang X, Clark AJ, St John
            Smith E, Bennett DL
  TITLE     A humanized chemogenetic system inhibits murine pain-related 
            behavior and hyperactivity in human sensory neurons.
  JOURNAL   Sci Transl Med. 2023 Oct 4;15(716):eadh3839. doi: 
            10.1126/scitranslmed.adh3839. Epub 2023 Oct 4.
  PUBMED    37792955
REFERENCE   2  (bases 1 to 7237)
  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 Transl 
            Med."; date: "2023-10-4"; volume: "15(716):eadh3839. doi"; pages: " 
            10.1126/scitranslmed.adh3839. Epub 2023 Oct 4"
COMMENT     Sequence Label: pAAV-CAG-mCherry-Tandem-PSAM4-GlyR
FEATURES             Location/Qualifiers
     source          1..7237
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        225..604
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          882..1898
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1822..1844
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1906..1925
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     1953..1972
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1953..1971
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1997..2701
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(2139..2157)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2407..2426
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2723..2779
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2789..2842
                     /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             4145..4162
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4185..4202)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4191..4415
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4500..4640
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4500..4629
                     /label=AAV2 ITR
     rep_origin      4715..5170
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4802..4821)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5012..5033
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5187..5206)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5306..5328
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5366..5384)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5452..5556
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5557..5625,5626..6417)
                     /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(5775..5794)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6588..7176
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7077..7096
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggcaattc agtcgataac tataacggtc ctaaggtagc gatttaaata
      181 cgcgctctct taaggtagcc ccgggacgcg tcaattgaga tctcgacatt gattattgac
      241 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
      301 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      361 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      421 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      481 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      541 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      601 catggtcgag gtgagcccca cgttctgctt cactctcccc atctcccccc cctccccacc
      661 cccaattttg tatttattta ttttttaatt attttgtgca gcgatggggg cggggggggg
      721 gggggggsgs gccvggsggg gggggggggg gggggggggg gggsggggsg rggcggagag
      781 gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt tccttttatg gcgaggcggc
      841 ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc gggagtcgct gcgcgctgcc
      901 ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc gccccggctc tgactgaccg
      961 cgttactccc acaggtgagc gggcgggacg gcccttctcc tccgggctgt aattagcgct
     1021 tggtttaatg acggcttgtt tcttttctgt ggctgcgtga aagccttgag gggctccggg
     1081 agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg cgtgtgtgtg tgcgtgggga
     1141 gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg ctgcgggcgc ggcgcggggc
     1201 tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg gggcggtgcc ccgcggtgcg
     1261 gggggggctg cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg gggtgagcag
     1321 ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca cccccctccc cgagttgctg
     1381 agcacggccc ggcttcgggt gcggggctcc gtacggggcg tggcgcgggg ctcgccgtgc
     1441 cgggcggggg gtggcggcag gtgggggtgc cgggcggggc ggggccgcct cgggccgggg
     1501 rgggctcggg ggrggggcgc ggcggccccc ggagcgccgg cggctgtcga ggcgcggcga
     1561 gccgcagcca ttgcctttta tggtaatcgt gcgagagggc gcagggactt cctttgtccc
     1621 aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac cccctctagc gggcgcgggg
     1681 cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga gggccttcgt gcgtcgccgc
     1741 gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac ggctgccttc
     1801 gggggggacg gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct
     1861 ctgctaacca tgttcatgcc ttcttctttt tcctacagct cctgggcaac gtgctggtta
     1921 ttgtgctgtc tcatcatttt ggcaaagaat tctaatacga ctcactatag ggagacccaa
     1981 gctggctagc gccaccatga gcaagggcga ggaggataac atggccatca tcaaggagtt
     2041 catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg
     2101 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
     2161 tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc
     2221 ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt
     2281 caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc
     2341 ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc
     2401 cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta
     2461 ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg
     2521 ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg
     2581 cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt
     2641 ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa
     2701 gaggcgcaaa aggggatccg gagccacgaa cttctctctg ttaaagcaag caggagacgt
     2761 ggaagaaaac cccggtcctg gatccggaga gggcagagga agtcttctaa catgcggtga
     2821 cgtggaggag aatcccggcc ctatgcgctg ttctccaggc ggcgtgtggc tcgccctggc
     2881 tgcttccctt ctgcacgtta gcctgcaggg tgagttccag cgcaaactgt ataaggagct
     2941 tgttaagaat tataaccccc tggagcggcc ggtcgcaaat gattcccagc cactgacagt
     3001 gtacttcagc ctctccttgc tgcagatcat ggacgtggat gaaaagaacc aggtgctgac
     3061 cactaatatt tggttgcaga tgtcctggac cgatcactac ttgcagtgga atgtgagcga
     3121 atacccaggt gtaaagactg taagattccc tgacggccaa atctggaaac cagatatcct
     3181 gctgtacaac agcgcagacg aaaggtttga tgcaacattt cacaccaacg tgggagtcaa
     3241 ttcttcaggc cactgcctgt acctgccccc tggaatcttc aagtcctcat gctatatcga
     3301 cgtccgctgg tttcccttcg acgtccagca ctgcaaactc aaattcggga gctggagcta
     3361 cggcggatgg agcctggatc tgcaaatgca ggaggctgac atctctggtt acatcccgaa
     3421 tggggagtgg gaccttgtgg gaatccccgg taaaagaagc gagcgatttt atgaatgctg
     3481 caaagagccc ttcccagatg tcaccttcac agtgaccatg cggagacgca tgggttatta
     3541 tctgatccaa atgtatatcc caagcttgct tatagtgatt ttgtcatgga tctccttctg
     3601 gattaatatg gacgccgctc cagctagggt cggactgggc atcaccacag tgctgacaat
     3661 gactactcag agctcaggca gccgagccag cttgcccaag gtttcttacg tgaaggccat
     3721 cgatatctgg atggctgtct gccttctgtt tgtcttcagc gcactgctgg aatacgccgc
     3781 tgtcaatttt gtgtctcgac agcataaaga gctgttgcgg ttcagaagaa aacgacgcca
     3841 ccacaaagag gatgaggcag gagaaggacg cttcaacttt agcgcctatg gtatgggacc
     3901 tgcttgcctc caggctaaag acggaatttc cgtgaaggga gccaacaata gcaacacaac
     3961 caacccaccc cctgctccat ctaagagccc ggaggaaatg cgcaaactct ttattcagag
     4021 agcgaaaaag atcgacaaaa tctcccggat cggattcccc atggctttcc tgattttcaa
     4081 catgttttat tggatcatct acaagattgt gcgaagggag gacgtacaca accagtaaac
     4141 cggtcatcat caccatcacc attgagttta aacccgctga tcagcctcga ctgtgccttc
     4201 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     4261 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
     4321 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
     4381 tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagatc
     4441 ctctcttaag gtagcatcga gatttaaatt agggataaca gggtaatggc gcgggccgca
     4501 ggaaccccta gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc
     4561 cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg
     4621 agcgcgcagc tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg
     4681 tatttcacac cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc
     4741 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cttagcgccc
     4801 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     4861 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     4921 aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     4981 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     5041 ctcaactcta tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat
     5101 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     5161 tttacaattt tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     5221 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
     5281 gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca
     5341 tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc
     5401 atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     5461 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     5521 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     5581 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     5641 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     5701 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     5761 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     5821 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     5881 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     5941 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     6001 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     6061 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     6121 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     6181 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     6241 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     6301 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     6361 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     6421 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     6481 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     6541 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     6601 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     6661 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     6721 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     6781 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     6841 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     6901 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     6961 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     7021 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     7081 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     7141 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     7201 cggttcctgg ccttttgctg gccttttgct cacatgt
//
