LOCUS       sequence-199198-        8146 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gi-coupled hM4D-IRES-mCitrine under the control of CaMKIIa promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8146)
  AUTHORS   Bryan Roth
  TITLE     Roth lab DREADDs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8146)
  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            "Unpublished"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-CaMKIIa-HA-hM4D(Gi)-IRES-mCitrine
FEATURES             Location/Qualifiers
     source          1..8146
                     /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        149..1437
                     /label=CaMKII promoter
                     /note="promoter for the mouse calcium/calmodulin-dependent 
                     kinase II alpha subunit"
     CDS             1467..1493
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    2949..3522
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3116..3133)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3343..3362
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     regulatory      3523..3532
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3529..3531,3532..3534,3535..4245)
                     /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(3574..3595)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3835..3854)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4182..4203
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    4273..4861
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4326..4346)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4744..4755)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4893..5369
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(5092..5111)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5409..5549
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5409..5538
                     /label=AAV2 ITR
     rep_origin      5624..6079
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5711..5730)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5921..5942
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6096..6115)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6215..6237
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6275..6293)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6361..6465
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6466..6534,6535..7326)
                     /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(6684..6703)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7497..8085
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7986..8005
                     /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 gcgtttaaca ttatggcctt aggtcacttc atctccatgg
      181 ggttcttctt ctgattttct agaaaatgag atgggggtgc agagagcttc ctcagtgacc
      241 tgcccagggt cacatcagaa atgtcagagc tagaacttga actcagatta ctaatcttaa
      301 attccatgcc ttgggggcat gcaagtacga tatacagaag gagtgaactc attagggcag
      361 atgaccaatg agtttaggaa agaagagtcc agggcagggt acatctacac cacccgccca
      421 gccctgggtg agtccagcca cgttcacctc attatagttg cctctctcca gtcctacctt
      481 gacgggaagc acaagcagaa actgggacag gagccccagg agaccaaatc ttcatggtcc
      541 ctctgggagg atgggtgggg agagctgtgg cagaggcctc aggaggggcc ctgctgctca
      601 gtggtgacag ataggggtga gaaagcagac agagtcattc cgtcagcatt ctgggtctgt
      661 ttggtacttc ttctcacgct aaggtggcgg tgtgatatgc acaatggcta aaaagcaggg
      721 agagctggaa agaaacaagg acagagacag aggccaagtc aaccagacca attcccagag
      781 gaagcaaaga aaccattaca gagactacaa gggggaaggg aaggagagat gaattagctt
      841 cccctgtaaa ccttagaacc cagctgttgc cagggcaacg gggcaatacc tgtctcttca
      901 gaggagatga agttgccagg gtaactacat cctgtctttc tcaaggacca tcccagaatg
      961 tggcacccac tagccgttac catagcaact gcctctttgc cccacttaat cccatcccgt
     1021 ctgttaaaag ggccctatag ttggaggtgg gggaggtagg aagagcgatg atcacttgtg
     1081 gactaagttt gttcgcatcc ccttctccaa ccccctcagt acatcaccct gggggaacag
     1141 ggtccacttg ctcctgggcc cacacagtcc tgcagtattg tgtatataag gccagggcaa
     1201 agaggagcag gttttaaagt gaaaggcagg caggtgttgg ggaggcagtt accggggcaa
     1261 cgggaacagg gcgtttcgga ggtggttgcc atggggacct ggatgctgac gaaggctcgc
     1321 gaggctgtga gcagccacag tgccctgctc agaagcccca agctcgtcag tcaagccggt
     1381 tctccgtttg cactcaggag cacgggcagg cgagtggccc ctagttctgg gggcagctct
     1441 agagcggtac cggatccgcc accatgtacc catacgatgt tccagattac gctatggcca
     1501 acttcacacc tgtcaatggc agctcgggca atcagtccgt gcgcctggtc acgtcatcat
     1561 cccacaatcg ctatgagacg gtggaaatgg tcttcattgc cacagtgaca ggctccctga
     1621 gcctggtgac tgtcgtgggc aacatcctgg tgatgctgtc catcaaggtc aacaggcagc
     1681 tgcagacagt caacaactac ttcctcttca gcctggcgtg tgctgatctc atcataggcg
     1741 ccttctccat gaacctctac accgtgtaca tcatcaaggg ctactggccc ctgggcgccg
     1801 tggtctgcga cctgtggctg gccctggact gcgtggtgag caacgcctcc gtcatgaacc
     1861 ttctcatcat cagctttgac cgctacttct gcgtcaccaa gcctctcacc taccctgccc
     1921 ggcgcaccac caagatggca ggcctcatga ttgctgctgc ctgggtactg tccttcgtgc
     1981 tctgggcgcc tgccatcttg ttctggcagt ttgtggtggg taagcggacg gtgcccgaca
     2041 accagtgctt catccagttc ctgtccaacc cagcagtgac ctttggcaca gccattgctg
     2101 gcttctacct gcctgtggtc atcatgacgg tgctgtacat ccacatctcc ctggccagtc
     2161 gcagccgagt ccacaagcac cggcccgagg gcccgaagga gaagaaagcc aagacgctgg
     2221 ccttcctcaa gagcccacta atgaagcaga gcgtcaagaa gcccccgccc ggggaggccg
     2281 cccgggagga gctgcgcaat ggcaagctgg aggaggcccc cccgccagcg ctgccaccgc
     2341 caccgcgccc cgtggctgat aaggacactt ccaatgagtc cagctcaggc agtgccaccc
     2401 agaacaccaa ggaacgccca gccacagagc tgtccaccac agaggccacc acgcccgcca
     2461 tgcccgcccc tcccctgcag ccgcgggccc tcaacccagc ctccagatgg tccaagatcc
     2521 agattgtgac gaagcagaca ggcaatgagt gtgtgacagc cattgagatt gtgcctgcca
     2581 cgccggctgg catgcgccct gcggccaacg tggcccgcaa gttcgccagc atcgctcgca
     2641 accaggtgcg caagaagcgg cagatggcgg cccgggagcg caaagtgaca cgaacgatct
     2701 ttgccattct gcttgccttc atcctcacct ggacgcccta caacgtcatg gtcctggtga
     2761 acaccttctg ccagagctgc atccctgaca cggtgtggtc cattggctac tggctctgct
     2821 acgtcaacag caccatcaac cctgcctgct atgctctgtg caacgccacc tttaaaaaga
     2881 ccttccggca cctgctgctg tgccagtatc ggaacatcgg cactgccagg taggcggccg
     2941 cgatatcgcc cctctccctc ccccccccct aacgttactg gccgaagccg cttggaataa
     3001 ggccggtgtg cgtttgtcta tatgttattt tccaccatat tgccgtcttt tggcaatgtg
     3061 agggcccgga aacctggccc tgtcttcttg acgagcattc ctaggggtct ttcccctctc
     3121 gccaaaggaa tgcaaggtct gttgaatgtc gtgaaggaag cagttcctct ggaagcttct
     3181 tgaagacaaa caacgtctgt agcgaccctt tgcaggcagc ggaacccccc acctggcgac
     3241 aggtgcctct gcggccaaaa gccacgtgta taagatacac ctgcaaaggc ggcacaaccc
     3301 cagtgccacg ttgtgagttg gatagttgtg gaaagagtca aatggctctc ctcaagcgta
     3361 ttcaacaagg ggctgaagga tgcccagaag gtaccccatt gtatgggatc tgatctgggg
     3421 cctcggtgca catgctttac atgtgtttag tcgaggttaa aaaaacgtct aggccccccg
     3481 aaccacgggg acgtggtttt cctttgaaaa acacgatgat aagccaccat ggtgagcaag
     3541 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     3601 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     3661 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     3721 ttcggctacg gcctgatgtg cttcgcccgc taccccgacc acatgaagca gcacgacttc
     3781 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     3841 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     3901 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     3961 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     4021 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     4081 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagctac
     4141 cagtccaaac tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     4201 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtgaga attcgatatc
     4261 aagcttatcg ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     4321 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     4381 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt
     4441 tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac
     4501 gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg aactttcgct
     4561 ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca
     4621 ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt
     4681 ccttggctgc tcgcctatgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc
     4741 ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct
     4801 cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg
     4861 catcgatacc gagcgctgct cgagagatct acgggtggca tccctgtgac ccctccccag
     4921 tgcctctcct ggccctggaa gttgccactc cagtgcccac cagccttgtc ctaataaaat
     4981 taagttgcat cattttgtct gactaggtgt ccttctataa tattatgggg tggagggggg
     5041 tggtatggag caaggggcaa gttgggaaga caacctgtag ggcctgcggg gtctattggg
     5101 aaccaagctg gagtgcagtg gcacaatctt ggctcactgc aatctccgcc tcctgggttc
     5161 aagcgattct cctgcctcag cctcccgagt tgttgggatt ccaggcatgc atgaccaggc
     5221 tcagctaatt tttgtttttt tggtagagac ggggtttcac catattggcc aggctggtct
     5281 ccaactccta atctcaggtg atctacccac cttggcctcc caaattgctg ggattacagg
     5341 cgtgaaccac tgctcccttc cctgtccttc tgattttgta ggtaaccacg tgcggaccga
     5401 gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct
     5461 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt
     5521 gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca
     5581 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     5641 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     5701 ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     5761 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     5821 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     5881 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     5941 ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     6001 tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     6061 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     6121 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     6181 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     6241 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     6301 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     6361 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     6421 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     6481 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     6541 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     6601 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     6661 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     6721 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     6781 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     6841 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     6901 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     6961 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     7021 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     7081 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     7141 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     7201 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     7261 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     7321 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     7381 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     7441 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     7501 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     7561 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     7621 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     7681 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     7741 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     7801 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     7861 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     7921 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     7981 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     8041 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     8101 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt
//
