LOCUS       sequence-214300-        7929 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 7929)
  AUTHORS   Bryan Roth
  TITLE     Roth lab DREADDs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7929)
  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            "Unpublished"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..7929
                     /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        175..622
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(660..693)
                     /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(744..777)
                     /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(792..1502,1503..1505,1506..1508))
                     /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(834..855)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1183..1202
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1442..1463
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1505..1514
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    1515..2088
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1675..1694)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     1904..1921
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(3874..3900)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    3914..3947
                     /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    3998..4031
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4056..4644
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4109..4129)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4527..4538)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4676..5152
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4875..4894)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5192..5332
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5192..5321
                     /label=AAV2 ITR
     rep_origin      5407..5862
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5494..5513)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5704..5725
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5879..5898)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5998..6020
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        6144..6248
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6249..6317,6318..7109)
                     /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(6467..6486)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7280..7868
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7769..7788
                     /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 gcgtgtgtct agactgcaga gggccctgcg tatgagtgca
      181 agtgggtttt aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga
      241 cccactggac aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg
      301 gaggggaaac aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt
      361 gccttcgccc ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac
      421 gtcactcgcc ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc
      481 cgccggccca gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca
      541 tctgcgctgc ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt
      601 cgtgtcgtgc ctgagagcgc agtcgagaag gtaccggatc ctctagagtc gactccggaa
      661 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
      721 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
      781 gcgccctatc acttgtacag ctcgtccatg ccgagagtga tcccggcggc ggtcacgaac
      841 tccagcagga ccatgtgatc gcgcttctcg ttggggtctt tgctcagttt ggactggtag
      901 ctcaggtagt ggttgtcggg cagcagcacg gggccgtcgc cgatgggggt gttctgctgg
      961 tagtggtcgg cgagctgcac gctgccgtcc tcgatgttgt ggcggatctt gaagttcacc
     1021 ttgatgccgt tcttctgctt gtcggccatg atatagacgt tgtggctgtt gtagttgtac
     1081 tccagcttgt gccccaggat gttgccgtcc tccttgaagt cgatgccctt cagctcgatg
     1141 cggttcacca gggtgtcgcc ctcgaacttc acctcggcgc gggtcttgta gttgccgtcg
     1201 tccttgaaga agatggtgcg ctcctggacg tagccttcgg gcatggcgga cttgaagaag
     1261 tcgtgctgct tcatgtggtc ggggtagcgg gcgaagcaca tcaggccgta gccgaaggtg
     1321 gtcacgaggg tgggccaggg cacgggcagc ttgccggtgg tgcagatgaa cttcagggtc
     1381 agcttgccgt aggtggcatc gccctcgccc tcgccggaca cgctgaactt gtggccgttt
     1441 acgtcgccgt ccagctcgac caggatgggc accaccccgg tgaacagctc ctcgcccttg
     1501 ctcaccatgg tggcttatca tcgtgttttt caaaggaaaa ccacgtcccc gtggttcggg
     1561 gggcctagac gtttttttaa cctcgactaa acacatgtaa agcatgtgca ccgaggcccc
     1621 agatcagatc ccatacaatg gggtaccttc tgggcatcct tcagcccctt gttgaatacg
     1681 cttgaggaga gccatttgac tctttccaca actatccaac tcacaacgtg gcactggggt
     1741 tgtgccgcct ttgcaggtgt atcttataca cgtggctttt ggccgcagag gcacctgtcg
     1801 ccaggtgggg ggttccgctg cctgcaaagg gtcgctacag acgttgtttg tcttcaagaa
     1861 gcttccagag gaactgcttc cttcacgaca ttcaacagac cttgcattcc tttggcgaga
     1921 ggggaaagac ccctaggaat gctcgtcaag aagacagggc caggtttccg ggccctcaca
     1981 ttgccaaaag acggcaatat ggtggaaaat aacatataga caaacgcaca ccggccttat
     2041 tccaagcggc ttcggccagt aacgttaggg gggggggagg gagaggggcg atatcgcggc
     2101 cgcctacaag gcctgctcgg gtgcgcgctt gtgaaaaatg accgactgtc tctgctggta
     2161 ctgctgcttg cgcctctttt ttttgtcaca ctggcacagc agcagcatct tgaaagtggt
     2221 tctgaatgtt ttgttgcaca gagcatagca cacggggttc acggtgctgt tgatgtagca
     2281 cagccagtag cccagattcc aaaaggtttt gggtatgcag ctgtcacaaa aggtgttcac
     2341 cagaaccatg atgttgtatg gggtccaagt gatgatgaag gcaagcaaga tcgcactgag
     2401 ggtctgggcc gctttcttct ccttgaccag ggacatcctt ttccgcttag tgatctgact
     2461 tctggtcttc agagcaaacc tcttggccag agtggcttcc ttgaaggaca gaggtagagt
     2521 ggccgtgctc ttacccactg aggagttgac gtcagaagtc ttagctgtgt ccacggctga
     2581 ctctagctgg atgggaagct tggagaagct ttttggaaaa ctgcctccat cgtccacgct
     2641 cttctgggcc tgcagcttgt cggctttcct ctccaagtcc accatcccca gctcctcctc
     2701 aggcacctgc aggttgtccg atgagggtaa cttggtggag ttgaggatgg tgctgtgacc
     2761 cggaagcttg agcacgatgg agtagatggc tctcgtctcg gagccaatgt cctcctcgtc
     2821 ggaggaggcg gagttctcca gggaggcagc agcatcattg ttgttccaac tgtcactgct
     2881 gctgtggtct tggtccatct gctcggagct gggtttccag ctcttggttg tgaaccagaa
     2941 gtggcagcgg ccatacttcc tcctgttgga gcgtttcatg ctttgctgtt gaagttcgta
     3001 actgctgcag cttcgagaac tgcccgtggg gtggacaaag ttttctgtct ctgcctctgt
     3061 cccagaggct tgcaggccag caagctcttt ggtacgcttt tcagtttcct tatagatcct
     3121 ccagtataaa atagtcataa tggtgacagg catataaaaa ccagcgatgg ctgtgccaaa
     3181 agtaatggtg ggctcactga ggaactgaat gaagcactct cccggaggca cagttctctt
     3241 tccaacaaag tattgccaga acaagatggc aggagcccaa aggacaaagg agatgaccca
     3301 agccagaccg atcatcacac cggctctctt tgttgttcgt ttggctcggt acgtgagcgg
     3361 cctcgtgatg gaaaagtatc tgtcaaagct gatgaccaga agattcataa cagaggcatt
     3421 gctggctacg cagtcaatgg caagccagag gtcacaggcc aagttcccta aggcccatcg
     3481 attcatgatg atgtaggtcg taaacagatt cattgaaatg accccgataa tcagatcggc
     3541 acaggccagg cttaagagga agtagttgtt gaccgtcttc agctgcttgt tgaccttaaa
     3601 tgacacaatt accaggatgt tgccgatgat ggtcaccaag gccaggatgc ccgttaagaa
     3661 agcgatgaag accacttgcc agacggtatg acctcccaga gggtcatcgg tggtaccgtc
     3721 tggagaggag aaattgccag ctgctcgaga aacattgtag ctgccgaaat gagtgacggt
     3781 tcccgggggc agccctgcat cggaggggct gtgtatccag gaggagctga tgtttggaaa
     3841 caaaggcgag gttgtactgt tattgtgcaa ggtagcgtaa tctggaacat cgtatgggta
     3901 catggtggct agcataactt cgtataaagt atcctatacg aagttatttg ccttaaccca
     3961 gaaattatca ctgttattct ttagaatggt gcaaagaata acttcgtata atgtatgcta
     4021 tacgaagtta tgaattcgat atcaagctta tcgataatca acctctggat tacaaaattt
     4081 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     4141 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     4201 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     4261 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     4321 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     4381 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     4441 tgtcggggaa atcatcgtcc tttccttggc tgctcgccta tgttgccacc tggattctgc
     4501 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     4561 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
     4621 tctccctttg ggccgcctcc ccgcatcgat accgagcgct gctcgagaga tctacgggtg
     4681 gcatccctgt gacccctccc cagtgcctct cctggccctg gaagttgcca ctccagtgcc
     4741 caccagcctt gtcctaataa aattaagttg catcattttg tctgactagg tgtccttcta
     4801 taatattatg gggtggaggg gggtggtatg gagcaagggg caagttggga agacaacctg
     4861 tagggcctgc ggggtctatt gggaaccaag ctggagtgca gtggcacaat cttggctcac
     4921 tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct cagcctcccg agttgttggg
     4981 attccaggca tgcatgacca ggctcagcta atttttgttt ttttggtaga gacggggttt
     5041 caccatattg gccaggctgg tctccaactc ctaatctcag gtgatctacc caccttggcc
     5101 tcccaaattg ctgggattac aggcgtgaac cactgctccc ttccctgtcc ttctgatttt
     5161 gtaggtaacc acgtgcggac cgagcggccg caggaacccc tagtgatgga gttggccact
     5221 ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg
     5281 ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg
     5341 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac
     5401 catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     5461 tgaccgctac acttgccagc gccttagcgc ccgctccttt cgctttcttc ccttcctttc
     5521 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     5581 gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta
     5641 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     5701 atagtggact cttgttccaa actggaacaa cactcaactc tatctcgggc tattcttttg
     5761 atttataagg gattttgccg atttcggtct attggttaaa aaatgagctg atttaacaaa
     5821 aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta
     5881 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     5941 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     6001 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc
     6061 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag
     6121 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     6181 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     6241 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     6301 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     6361 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     6421 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     6481 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     6541 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     6601 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     6661 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     6721 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     6781 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     6841 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     6901 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     6961 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     7021 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     7081 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     7141 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     7201 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     7261 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     7321 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     7381 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     7441 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     7501 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     7561 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     7621 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     7681 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     7741 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     7801 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     7861 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     7921 ctcacatgt
//
