LOCUS       sequence-75399-e        7937 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Double floxed Gq-coupled hM3D-IRES-mCitrine under the control of 
            human synapsin promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7937)
  AUTHORS   Bryan Roth
  TITLE     Roth lab DREADDs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7937)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pAAV-hSyn-DIO-HA-hM3D(Gq)-IRES-mCitrine
FEATURES             Location/Qualifiers
     source          1..7937
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        186..633
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(671..704)
                     /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(755..788)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(800..1510,1511..1513,1514..1516))
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(842..863)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1191..1210
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1450..1471
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1513..1522
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    1523..2096
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1683..1702)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     1912..1929
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(3882..3908)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    3922..3955
                     /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    4006..4039
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4064..4652
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4117..4137)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4535..4546)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4684..5160
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4883..4902)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5200..5340
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5200..5329
                     /label=AAV2 ITR
     rep_origin      5415..5870
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5502..5521)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5712..5733
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5887..5906)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6006..6028
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        6152..6256
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6257..6325,6326..7117)
                     /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(6475..6494)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7288..7876
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7777..7796
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtgtgtc tagactgcag agggccctgc
      181 gtatgagtgc aagtgggttt taggaccagg atgaggcggg gtgggggtgc ctacctgacg
      241 accgaccccg acccactgga caagcaccca acccccattc cccaaattgc gcatccccta
      301 tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt gcgcactgcc agcttcagca
      361 ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc accgccgcct cagcactgaa
      421 ggcgcgctga cgtcactcgc cggtcccccg caaactcccc ttcccggcca ccttggtcgc
      481 gtccgcgccg ccgccggccc agccggaccg caccacgcga ggcgcgagat aggggggcac
      541 gggcgcgacc atctgcgctg cggcgccggc gactcagcgc tgcctcagtc tgcggtgggc
      601 agcggaggag tcgtgtcgtg cctgagagcg cagtcgagaa ggtaccggat cctctagagt
      661 cgactccgga ataacttcgt ataggatact ttatacgaag ttatgcagaa tggtagctgg
      721 attgtagctg ctattagcaa tatgaaacct cttaataact tcgtatagca tacattatac
      781 gaagttatgg cgcgcctcac ttgtacagct cgtccatgcc gagagtgatc ccggcggcgg
      841 tcacgaactc cagcaggacc atgtgatcgc gcttctcgtt ggggtctttg ctcagtttgg
      901 actggtagct caggtagtgg ttgtcgggca gcagcacggg gccgtcgccg atgggggtgt
      961 tctgctggta gtggtcggcg agctgcacgc tgccgtcctc gatgttgtgg cggatcttga
     1021 agttcacctt gatgccgttc ttctgcttgt cggccatgat atagacgttg tggctgttgt
     1081 agttgtactc cagcttgtgc cccaggatgt tgccgtcctc cttgaagtcg atgcccttca
     1141 gctcgatgcg gttcaccagg gtgtcgccct cgaacttcac ctcggcgcgg gtcttgtagt
     1201 tgccgtcgtc cttgaagaag atggtgcgct cctggacgta gccttcgggc atggcggact
     1261 tgaagaagtc gtgctgcttc atgtggtcgg ggtagcgggc gaagcacatc aggccgtagc
     1321 cgaaggtggt cacgagggtg ggccagggca cgggcagctt gccggtggtg cagatgaact
     1381 tcagggtcag cttgccgtag gtggcatcgc cctcgccctc gccggacacg ctgaacttgt
     1441 ggccgtttac gtcgccgtcc agctcgacca ggatgggcac caccccggtg aacagctcct
     1501 cgcccttgct caccatggtg gcttatcatc gtgtttttca aaggaaaacc acgtccccgt
     1561 ggttcggggg gcctagacgt ttttttaacc tcgactaaac acatgtaaag catgtgcacc
     1621 gaggccccag atcagatccc atacaatggg gtaccttctg ggcatccttc agccccttgt
     1681 tgaatacgct tgaggagagc catttgactc tttccacaac tatccaactc acaacgtggc
     1741 actggggttg tgccgccttt gcaggtgtat cttatacacg tggcttttgg ccgcagaggc
     1801 acctgtcgcc aggtgggggg ttccgctgcc tgcaaagggt cgctacagac gttgtttgtc
     1861 ttcaagaagc ttccagagga actgcttcct tcacgacatt caacagacct tgcattcctt
     1921 tggcgagagg ggaaagaccc ctaggaatgc tcgtcaagaa gacagggcca ggtttccggg
     1981 ccctcacatt gccaaaagac ggcaatatgg tggaaaataa catatagaca aacgcacacc
     2041 ggccttattc caagcggctt cggccagtaa cgttaggggg gggggaggga gaggggcgat
     2101 atcgcggccg cctacaaggc ctgctcgggt gcgcgcttgt gaaaaatgac cgactgtctc
     2161 tgctggtact gctgcttgcg cctctttttt ttgtcacact ggcacagcag cagcatcttg
     2221 aaagtggttc tgaatgtttt gttgcacaga gcatagcaca cggggttcac ggtgctgttg
     2281 atgtagcaca gccagtagcc cagattccaa aaggttttgg gtatgcagct gtcacaaaag
     2341 gtgttcacca gaaccatgat gttgtatggg gtccaagtga tgatgaaggc aagcaagatc
     2401 gcactgaggg tctgggccgc tttcttctcc ttgaccaggg acatcctttt ccgcttagtg
     2461 atctgacttc tggtcttcag agcaaacctc ttggccagag tggcttcctt gaaggacaga
     2521 ggtagagtgg ccgtgctctt acccactgag gagttgacgt cagaagtctt agctgtgtcc
     2581 acggctgact ctagctggat gggaagcttg gagaagcttt ttggaaaact gcctccatcg
     2641 tccacgctct tctgggcctg cagcttgtcg gctttcctct ccaagtccac catccccagc
     2701 tcctcctcag gcacctgcag gttgtccgat gagggtaact tggtggagtt gaggatggtg
     2761 ctgtgacccg gaagcttgag cacgatggag tagatggctc tcgtctcgga gccaatgtcc
     2821 tcctcgtcgg aggaggcgga gttctccagg gaggcagcag catcattgtt gttccaactg
     2881 tcactgctgc tgtggtcttg gtccatctgc tcggagctgg gtttccagct cttggttgtg
     2941 aaccagaagt ggcagcggcc atacttcctc ctgttggagc gtttcatgct ttgctgttga
     3001 agttcgtaac tgctgcagct tcgagaactg cccgtggggt ggacaaagtt ttctgtctct
     3061 gcctctgtcc cagaggcttg caggccagca agctctttgg tacgcttttc agtttcctta
     3121 tagatcctcc agtataaaat agtcataatg gtgacaggca tataaaaacc agcgatggct
     3181 gtgccaaaag taatggtggg ctcactgagg aactgaatga agcactctcc cggaggcaca
     3241 gttctctttc caacaaagta ttgccagaac aagatggcag gagcccaaag gacaaaggag
     3301 atgacccaag ccagaccgat catcacaccg gctctctttg ttgttcgttt ggctcggtac
     3361 gtgagcggcc tcgtgatgga aaagtatctg tcaaagctga tgaccagaag attcataaca
     3421 gaggcattgc tggctacgca gtcaatggca agccagaggt cacaggccaa gttccctaag
     3481 gcccatcgat tcatgatgat gtaggtcgta aacagattca ttgaaatgac cccgataatc
     3541 agatcggcac aggccaggct taagaggaag tagttgttga ccgtcttcag ctgcttgttg
     3601 accttaaatg acacaattac caggatgttg ccgatgatgg tcaccaaggc caggatgccc
     3661 gttaagaaag cgatgaagac cacttgccag acggtatgac ctcccagagg gtcatcggtg
     3721 gtaccgtctg gagaggagaa attgccagct gctcgagaaa cattgtagct gccgaaatga
     3781 gtgacggttc ccgggggcag ccctgcatcg gaggggctgt gtatccagga ggagctgatg
     3841 tttggaaaca aaggcgaggt tgtactgtta ttgtgcaagg tagcgtaatc tggaacatcg
     3901 tatgggtaca tggtggctag cataacttcg tataaagtat cctatacgaa gttatttgcc
     3961 ttaacccaga aattatcact gttattcttt agaatggtgc aaagaataac ttcgtataat
     4021 gtatgctata cgaagttatg aattcgatat caagcttatc gataatcaac ctctggatta
     4081 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     4141 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     4201 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     4261 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     4321 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     4381 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     4441 cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctatg ttgccacctg
     4501 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     4561 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac
     4621 gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgagcgctgc tcgagagatc
     4681 tacgggtggc atccctgtga cccctcccca gtgcctctcc tggccctgga agttgccact
     4741 ccagtgccca ccagccttgt cctaataaaa ttaagttgca tcattttgtc tgactaggtg
     4801 tccttctata atattatggg gtggaggggg gtggtatgga gcaaggggca agttgggaag
     4861 acaacctgta gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt ggcacaatct
     4921 tggctcactg caatctccgc ctcctgggtt caagcgattc tcctgcctca gcctcccgag
     4981 ttgttgggat tccaggcatg catgaccagg ctcagctaat ttttgttttt ttggtagaga
     5041 cggggtttca ccatattggc caggctggtc tccaactcct aatctcaggt gatctaccca
     5101 ccttggcctc ccaaattgct gggattacag gcgtgaacca ctgctccctt ccctgtcctt
     5161 ctgattttgt aggtaaccac gtgcggaccg agcggccgca ggaaccccta gtgatggagt
     5221 tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc
     5281 gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg
     5341 ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc
     5401 aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     5461 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     5521 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     5581 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg
     5641 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     5701 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcgggcta
     5761 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     5821 ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac
     5881 tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc
     5941 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac
     6001 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg
     6061 aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta
     6121 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     6181 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     6241 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     6301 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     6361 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     6421 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     6481 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     6541 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     6601 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     6661 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     6721 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     6781 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     6841 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     6901 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     6961 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     7021 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     7081 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     7141 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     7201 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     7261 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     7321 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     7381 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
     7441 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     7501 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     7561 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     7621 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     7681 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     7741 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     7801 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     7861 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     7921 gccttttgct cacatgt
//
