LOCUS       sequence-432468-        6951 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated, Cre-dependent co-expression of GtACR2 and 
            soma-targeted Voltron2 (ORCHID) for assessing inhibitory receptor 
            driving force through voltage imaging with concurrent activation of 
            GtACR2..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6951)
  AUTHORS   Selfe JS, Steyn TJS, Shorer EF, Burman RJ, Dusterwald KM, Kraitzick 
            AZ, Abdelfattah AS, Schreiter ER, Newey SE, Akerman CJ, Raimondo JV
  TITLE     All-optical reporting of inhibitory receptor driving force in the 
            nervous system.
  JOURNAL   Nat Commun. 2024 Oct 16;15(1):8913. doi: 10.1038/s41467-024-53074-y.
  PUBMED    39414774
REFERENCE   2  (bases 1 to 6951)
  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: "Nat 
            Commun."; date: "2024-10-16"; volume: "15(1):8913. doi"; pages: " 
            10.1038/s41467-024-53074-y"
COMMENT     Sequence Label: pAAV-EF1-alpha-DIO-GtACR2-P2A-Voltron2ST-W3SL
FEATURES             Location/Qualifiers
     source          1..6951
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        177..388
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             401..669
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     protein_bind    complement(703..736)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(787..820)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2696..2752)
                     /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"
     regulatory      3631..3640
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3647..3680
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    3731..3764
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(3834..3854)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4214..4354
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4214..4343
                     /label=AAV2 ITR
     rep_origin      4429..4884
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4516..4535)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4726..4747
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4901..4920)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5020..5042
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5080..5098)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5166..5270
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5271..5339,5340..6131)
                     /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(5489..5508)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6302..6890
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6791..6810
                     /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 gcggccgcac gcgtaaggat ctgcgatcgc tccggtgccc gtcagtgggc
      181 agagcgcaca tcgcccacag tccccgagaa gttgggggga ggggtcggca attgaacggg
      241 tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct
      301 ttttcccgag ggtgggggag aaccgtatat aagtgcagta gtcgccgtga acgttctttt
      361 tcgcaacggg tttgccgcca gaacacagct gaagcttcga ggggctcgca tctctccttc
      421 acgcgcccgc cgccctacct gaggccgcca tccacgccgg ttgagtcgcg ttctgccgcc
      481 tcccgcctgt ggtgcctcct gaactgcgtc cgccgtctag gtaagtttaa agctcaggtc
      541 gagaccgggc ctttgtccgg cgctcccttg gagcctacct agactcagcc ggctctccac
      601 gctttgcctg accctgcttg ctcaactcta cgtctttgtt tcgttttctg ttctgcgccg
      661 ttacagatcc aagctgtgac cggcgcctac gtcgactccg gaataacttc gtataggata
      721 ctttatacga agttatgcag aatggtagct ggattgtagc tgctattagc aatatgaaac
      781 ctcttaataa cttcgtatag catacattat acgaagttat ggcgcgtaat tacttgtaca
      841 attagaatcg agtagcctcg gggaaatctg ttgcgcaaga gatgaagcta tcaatagatg
      901 acatactcct catgtcaatc actccttcag tcctggcagg gaggggagcc acgggacttg
      961 gcatactgga catctcaagc tcttcgggag gcttagactg tggagccatt tcctttgtgt
     1021 tcaggatagg ctgactttgc acctcgttct cgtagcagaa cacgttgatg tcgatctggt
     1081 ccagggggat gtactcgccc tcgctggtga tcctgctctt agtggttggc tcgccggaaa
     1141 tctcgagcgt cgacagccag cgcgcgatct cgctgccgat caggtccggg ttgtcttctt
     1201 gcagcagatt cagacccggg ccgatgtcca cagccttgca gttaggcagg cttttggcca
     1261 ggcgagcggc ttcggccggt gggatcagaa cgcctggggt gccccagaac agcagcttcg
     1321 ggacagggga ctggtgcagc cagtccatgt attcttcgac cagcgcgacg atgttcgctg
     1381 gctcaccggc gattggcagc tcgtttggga agcgccacag tggctcgcgg tcaacaggat
     1441 tcaggaacgg ctcgcggtaa tggtccatct cgacttcagt cagcgggcgg acgacaccca
     1501 tcggcagcgt accctcgata aaaacgttct gatcgatgat cagcttgcgg ccgacgtcgg
     1561 tggtgcggaa ggcctggaag gtctcgcggg caaattctgg ccattcgtcc caggtcggga
     1621 tagggtggat gaactccata aatgcaatac ctttgacgcg ctctggattg cgcttggccc
     1681 agtggaaacc cagagcggag ccccagtcgt gaatgaccag gacgacctct tccagaccca
     1741 gggcttcgat gaaggcatcc atgaagcgga cgtggtcgtc gaagaaataa cccaggtctg
     1801 gtttgtcgga tttgcccata ccgatcaggt ctggagcaat gcagcgatgg gtcggtgcaa
     1861 catgcgggat gatgttgcgc cacacgtagg aggaggtcgg gttaccgtgc aggaacagca
     1921 caggggtgcc atcgcgcgga ccaacatcga cgtagtgcat gcgctcgccc aggacttcca
     1981 cataatgggg gtcgaatgga aagccggtac cgatcttgaa gatagtctca tgggcaatga
     2041 ggcaccccat tccaaacagt ggcttagcga gcacatccag gattcccatg agaatggcct
     2101 cattgtcgac ggacagcacg ccactaccag acccgaaagc ccacacgaca ggataaaaca
     2161 cccagaggat tgacaggaag acgagcagac tgacatagga gctatggtac tcggggcttt
     2221 tcttcagatc gtctttccaa gtggcgttat acatcccgta tgtcaggact gcctgcatca
     2281 cacatccagc gataaagtac acccatttga acttgtgttc atcctcgaag tcgcccagaa
     2341 taccaaacac gatcatgaaa atgtctgcgc cgatgaccca agaaatcatc actccgccca
     2401 tcttggtcat gacgatgaga tcgagcagga gcagtggtgt ggtcagcacc cagttaatat
     2461 atcttgcgta gaccacctgc ctttcaccat tcattgtcag gtcctggtag ttcacggcca
     2521 ttgcatagta agcggtagcg gcgatagtgc agatggcaat gttgatatag tatgcgctct
     2581 tcagctctga aggccgaaag tgcacagcca ctccaaaagc aatagcagca gcagccataa
     2641 tagcaaagac aatccactgt gcaatcatgc cggtctcggt ttccacgtca gccatgggac
     2701 cggggttttc ttccacgtct cctgcttgct ttaacagaga gaagttcgtg gctccggagc
     2761 ccctgccgaa cattctgatg ttgacctcgt cgttgggggg ggcattcttc tcgtggttca
     2821 gtccgaaggg gtcgtcggcc tcaggcttct cctggccagg gaggcaagcc gggtcccact
     2881 ttccctcgag cactccccag gcggtggacc acaggatcac gccgtaggtg ttcttgcaca
     2941 ggccgtcggc gagcagatac aggatggcgc tcacgttctc gtggatcacg cacacgccgg
     3001 tgggggaaat cagccacacg atggggaaca tggtccaggc gatgaagaag atcagcctca
     3061 gcatcttgat cctcaccacc accttctggg ccagaggggt gcccacggac tcgaagccgt
     3121 acagtcccac ctggaagatg gtgaacacca cgctgtattc gaagaccagg caggtagcgc
     3181 cgaagaagaa gaacagccac ttcatgtact cggcggggga gatggtagcg gtaatgccca
     3241 tcaccaccct gataatgctg gcggcctggg cgatggggtt cagagggatg ctcttgtact
     3301 tcaccagagc catgttggag atctggccca gcataatggg gcatgtgcac agccagctgg
     3361 ccatcctgga ccagggaaag aacctgccgt ccctcatctg cagtgtgcca ttgccggtgc
     3421 tggccagggc gtatgtgatg gactccacga agggcaggta cacgctctcc cagggcacct
     3481 tgggggccct gctgatcatg aagatggtct ggcatccgct caccacggcc cacaccagct
     3541 gcagcaggct cacgaaggtg ctgtcgatcc tgctcatgtt gtagcactct gtgtgggtgc
     3601 tggcccactc gccgtacacg acctgggagg ccatggtggc gctagcataa cttcgtataa
     3661 agtatcctat acgaagttat ttgccttaac ccagaaatta tcactgttat tctttagaat
     3721 ggtgcaaaga ataacttcgt ataatgtatg ctatacgaag ttatgaattc cgctcgagat
     3781 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     3841 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     3901 atggctttca ttttctcctc cttgtataaa tcctggttag ttcttgccac ggcggaactc
     3961 atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc
     4021 gtggtgttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat ctagctttat
     4081 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     4141 taacaacaac aattgcattc attttatgtt tcaggttcag ggggagatgt gggaggtttt
     4201 ttaaagcggc cgcaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc
     4261 tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc
     4321 tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc tgatgcggta ttttctcctt
     4381 acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac gcgccctgta
     4441 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     4501 gcgccttagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     4561 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     4621 acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca tcgccctgat
     4681 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     4741 aaactggaac aacactcaac tctatctcgg gctattcttt tgatttataa gggattttgc
     4801 cgatttcggt ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     4861 acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc tctgatgccg
     4921 catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
     4981 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     5041 ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt
     5101 tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa
     5161 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     5221 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     5281 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     5341 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     5401 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     5461 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     5521 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     5581 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     5641 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     5701 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     5761 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     5821 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     5881 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     5941 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     6001 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     6061 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     6121 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     6181 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     6241 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     6301 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     6361 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     6421 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     6481 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     6541 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     6601 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     6661 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     6721 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     6781 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     6841 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     6901 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg t
//
