LOCUS       sequence-441218-        7319 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Neuron-specific, Cre-dependent co-expression of FLAG-tagged 
            inhibitory KORD DREADD receptor, and 3x HA-tagged excitatory 
            hM3D(Gq) DREADD receptor..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7319)
  AUTHORS   Szelenyi ER, Navarrete JS, Murry AD, Zhang Y, Girven KS, Kuo L, 
            Cline MM, Bernstein MX, Burdyniuk M, Bowler B, Goodwin NL, Juarez B,
            Zweifel LS, Golden SA
  TITLE     An arginine-rich nuclear localization signal (ArgiNLS) strategy for 
            streamlined image segmentation of single cells.
  JOURNAL   Proc Natl Acad Sci U S A. 2024 Aug 6;121(32):e2320250121. doi: 
            10.1073/pnas.2320250121. Epub 2024 Jul 29.
  PUBMED    39074275
REFERENCE   2  (bases 1 to 7319)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2024-08-6"; volume: "121(32):e2320250121. 
            doi"; pages: " 10.1073/pnas.2320250121. Epub 2024 Jul 29"
COMMENT     Sequence Label: pAAV-hSyn-DIO-FLAG-KORD-P2A-3xHA-hM3D(Gq)
FEATURES             Location/Qualifiers
     source          1..7319
                     /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        145..592
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(628..661)
                     /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(712..745)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2545..2571)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2575..2601)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2605..2631)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2647..2703)
                     /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             3208..3219
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             complement(3928..3951)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      3951..3960
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3967..4000
                     /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    4051..4084
                     /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(4181..4201)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4583..4723
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4583..4712
                     /label=AAV2 ITR
     rep_origin      4798..5253
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4885..4904)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5095..5116
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5270..5289)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5389..5411
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        5535..5639
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5640..5708,5709..6500)
                     /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(5858..5877)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6671..7258
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7159..7178
                     /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 gcgtagtgca agtgggtttt aggaccagga tgaggcgggg
      181 tgggggtgcc tacctgacga ccgaccccga cccactggac aagcacccaa cccccattcc
      241 ccaaattgcg catcccctat cagagagggg gaggggaaac aggatgcggc gaggcgcgtg
      301 cgcactgcca gcttcagcac cgcggacagt gccttcgccc ccgcctggcg gcgcgcgcca
      361 ccgccgcctc agcactgaag gcgcgctgac gtcactcgcc ggtcccccgc aaactcccct
      421 tcccggccac cttggtcgcg tccgcgccgc cgccggccca gccggaccgc accacgcgag
      481 gcgcgagata ggggggcacg ggcgcgacca tctgcgctgc ggcgccggcg actcagcgct
      541 gcctcagtct gcggtgggca gcggaggagt cgtgtcgtgc ctgagagcgc agggtaccgt
      601 accggatcct ctagagtcga ctccggaata acttcgtata ggatacttta tacgaagtta
      661 tgcagaatgg tagctggatt gtagctgcta ttagcaatat gaaacctctt aataacttcg
      721 tatagcatac attatacgaa gttatggcgc gccttaggtg gcgaccgggg gatccttcaa
      781 ggcctgctcg ggtgcgcgct tgtgaaaaat gaccgactgt ctctgctggt actgctgctt
      841 gcgcctcttt tttttgtcac actggcacag cagcagcatc ttgaaagtgg ttctgaatgt
      901 tttgttgcac agagcatagc acacggggtt cacggtgctg ttgatgtagc acagccagta
      961 gcccagattc caaaaggttt tgggtatgca gctgtcacaa aaggtgttca ccagaaccat
     1021 gatgttgtat ggggtccaag tgatgatgaa ggcaagcaag atcgcactga gggtctgggc
     1081 cgctttcttc tccttgacca gggacatcct tttccgctta gtgatctgac ttctggtctt
     1141 cagagcaaac ctcttggcca gagtggcttc cttgaaggac agaggtagag tggccgtgct
     1201 cttacccact gaggagttga cgtcagaagt cttagctgtg tccacggctg actctagctg
     1261 gatgggaagc ttggagaagc tttttggaaa actgcctcca tcgtccacgc tcttctgggc
     1321 ctgcagcttg tcggctttcc tctccaagtc caccatcccc agctcctcct caggcacctg
     1381 caggttgtcc gatgagggta acttggtgga gttgaggatg gtgctgtgac ccggaagctt
     1441 gagcacgatg gagtagatgg ctctcgtctc ggagccaatg tcctcctcgt cggaggaggc
     1501 ggagttctcc agggaggcag cagcatcatt gttgttccaa ctgtcactgc tgctgtggtc
     1561 ttggtccatc tgctcggagc tgggtttcca gctcttggtt gtgaaccaga agtggcagcg
     1621 gccatacttc ctcctgttgg agcgtttcat gctttgctgt tgaagttcgt aactgctgca
     1681 gcttcgagaa ctgcccgtgg ggtggacaaa gttttctgtc tctgcctctg tcccagaggc
     1741 ttgcaggcca gcaagctctt tggtacgctt ttcagtttcc ttatagatcc tccagtataa
     1801 aatagtcata atggtgacag gcatataaaa accagcgatg gctgtgccaa aagtaatggt
     1861 gggctcactg aggaactgaa tgaagcactc tcccggaggc acagttctct ttccaacaaa
     1921 gtattgccag aacaagatgg caggagccca aaggacaaag gagatgaccc aagccagacc
     1981 gatcatcaca ccggctctct ttgttgttcg tttggctcgg tacgtgagcg gcctcgtgat
     2041 ggaaaagtat ctgtcaaagc tgatgaccag aagattcata acagaggcat tgctggctac
     2101 gcagtcaatg gcaagccaga ggtcacaggc caagttccct aaggcccatc gattcatgat
     2161 gatgtaggtc gtaaacagat tcattgaaat gaccccgata atcagatcgg cacaggccag
     2221 gcttaagagg aagtagttgt tgaccgtctt cagctgcttg ttgaccttaa atgacacaat
     2281 taccaggatg ttgccgatga tggtcaccaa ggccaggatg cccgttaaga aagcgatgaa
     2341 gaccacttgc cagacggtat gacctcccag agggtcatcg gtggtaccgt ctggagagga
     2401 gaaattgcca gctgctcgag aaacattgta gctgccgaaa tgagtgacgg ttcccggggg
     2461 cagccctgca tcggaggggc tgtgtatcca ggaggagctg atgtttggaa acaaaggcga
     2521 ggttgtactg ttattgtgca aggtagcgta gtctgggacg tcgtatgggt agccagcgta
     2581 gtctgggacg tcgtatgggt agccagcgta atccggaaca tcatacgggt atccttgttg
     2641 ccccatagga ccggggtttt cttccacgtc tcctgcttgc tttaacagag agaagttcgt
     2701 ggctccggat cctactggtt tattcatccc atcgatgtcc ctcaggtaag caggatcctg
     2761 aactgtattt cggactctgc tagtgctctg ccgctccatc ctcatcttca gtggaaagca
     2821 gaagtcccgg aaacaccgct tgaagttttc atcaagaaag gcgtagagaa tgggattcag
     2881 gctactgttg gtatagccta aggcgatgca gaagtaatag ctggagagag cagctgtgct
     2941 gtgggaggtg ctccccagag cctccaccag gatgaatatg tgaatgggag tccagcagac
     3001 gacgaagact gccaccacca ccaggaccag tctggtgatc ctacgcaggt tgcgatcttt
     3061 ctctcgggag ccagaaagga gccggacgct cttgagacgc aggatcatca gggtgtagca
     3121 gacgatgatg atgaggacag ggatcacgaa ggcaaagatg aagacgcaga tcttcatgaa
     3181 gaggtcccac caggagtagt catcatctgg gaactgcaag gagcactcaa tgacatcgac
     3241 gtcttccctg actttggtgc ctccaaggac tattgcagag atgccaacag atgacgacag
     3301 cagccagatg cagatattga tgatctttgc cttcaagggt gtgcggaagt ccaaagcctt
     3361 cacggggtgg cacacggcaa tgtagcggtc cacgctcatc atggtcaagg tgaagatgct
     3421 ggtgaacatg ttgtagtaat taatggaaat tactatcttg cacagcacat ccccaaaagg
     3481 ccaggaattc atcaagtaga ccgtactctg aaagggcatg gttgtagtaa ctaaagcatc
     3541 tgccaaagcc aggttaaata tgtaaatgtt ggttgctgtc ttcatctttg tgtatcggat
     3601 gatcacgaac atgaccagcg agttgcccac caagcccacg acgaacacta cggagtagac
     3661 cgccgtgatg atgaccggga tggccgggga gatgtgcgcg ggctccagct gcgcgtcctc
     3721 cgagccggcg ctgccgttgc tgtcgggctc ggcccagccg ggaaaccagg cgctgctgtt
     3781 ggggggcagg caggcgctcg gggcgcaggt agggcccggc tccccgcgga agatctgaat
     3841 cggggattca tttaaatctt cttctgataa taatttttgt tctaatgcat ctcgagtgga
     3901 tccgagctct gaattctcac cgatatcttt atcatcatca tctttataat ccatggtggc
     3961 gctagcataa cttcgtataa agtatcctat acgaagttat ttgccttaac ccagaaatta
     4021 tcactgttat tctttagaat ggtgcaaaga ataacttcgt ataatgtatg ctatacgaag
     4081 ttatgaattc gatatcaagc ttatcgatat cgaattccgc tcgagataat caacctctgg
     4141 attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat
     4201 gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt
     4261 tctcctcctt gtataaatcc tggttagttc ttgccacggc ggaactcatc gccgcctgcc
     4321 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgtttattt
     4381 gtgaaatttg tgatgctatt gctttatttg taaccatcta gctttatttg tgaaatttgt
     4441 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     4501 tgcattcatt ttatgtttca ggttcagggg gagatgtggg aggtttttta aagcggccgc
     4561 cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     4621 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     4681 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
     4741 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
     4801 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4861 cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4921 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     4981 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
     5041 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     5101 tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt gatttataag
     5161 ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     5221 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     5281 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     5341 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     5401 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     5461 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     5521 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     5581 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     5641 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5701 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     5761 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     5821 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     5881 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     5941 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     6001 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     6061 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     6121 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     6181 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     6241 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     6301 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     6361 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     6421 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     6481 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     6541 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     6601 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     6661 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     6721 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     6781 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     6841 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     6901 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     6961 taccggataa ggcgcagcgg tcgggctgaa cgggggttcg tgcacacagc ccagcttgga
     7021 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     7081 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     7141 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     7201 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     7261 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgt
//
