LOCUS       sequence-155887-        7148 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the DREADD receptor hM3Dq-mCherry under the control of 
            tet-inducible promoter for use in TetTaging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7148)
  AUTHORS   Zhang Z, Ferretti V, Guntan I, Moro A, Steinberg EA, Ye Z, Zecharia 
            AY, Yu X, Vyssotski AL, Brickley SG, Yustos R, Pillidge ZE, Harding 
            EC, Wisden W, Franks NP
  TITLE     Neuronal ensembles sufficient for recovery sleep and the sedative 
            actions of alpha2 adrenergic agonists.
  JOURNAL   Nat Neurosci. 2015 Apr;18(4):553-61. doi: 10.1038/nn.3957. Epub 2015
            Feb 23.
  PUBMED    25706476
REFERENCE   2  (bases 1 to 7148)
  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"; doi: "10.1038/nn.3957"; 
            journalName: "Nat Neurosci"; date: "2015-04"; volume: "18"; issue: 
            "4"; pages: "553-61"
COMMENT     Sequence Label: pAAV-PTRE-tight-hM3Dq-mCherry
FEATURES             Location/Qualifiers
     source          1..7148
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(30..52)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     152..171
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(188..643)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(325..346)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     537..556
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   718..858
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   729..858
                     /label=AAV2 ITR
     polyA_signal    898..1374
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     1156..1175
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     misc_feature    1406..1994
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1512..1523
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1921..1941
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    complement(2019..2052)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(2103..2136)
                     /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)."
     CDS             complement(2144..2854)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2422..2441)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2691..2709
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      2851..2860
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    4657..4690
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    4741..4774
                     /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)."
     promoter        complement(4814..5128)
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     primer_bind     complement(4817..4841)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    4888..4906
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4924..4942
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4959..4977
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4995..5013
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5031..5049
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5066..5084
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5102..5120
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     repeat_region   5139..5268
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(5330..5918)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5410..5429)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6089..6880,6881..6949))
                     /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     6712..6731
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6950..7054)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
       61 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
      121 gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt actgagagtg
      181 caccataaaa ttgtaaacgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
      241 agctcatttt ttaaccaata gaccgaaatc ggcaaaatcc cttataaatc aaaagaatag
      301 cccgagatag agttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
      361 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
      421 tcacccaaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
      481 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
      541 aagaaagcga aaggagcggg cgctaaggcg ctggcaagtg tagcggtcac gctgcgcgta
      601 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtactatgg ttgctttgac
      661 gtatgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgcccct
      721 gcaggcagct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg
      781 cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact
      841 ccatcactag gggttcctgc ggccgctcgg tccgcacgtg gttacctaca aaatcagaag
      901 gacagggaag ggagcagtgg ttcacgcctg taatcccagc aatttgggag gccaaggtgg
      961 gtagatcacc tgagattagg agttggagac cagcctggcc aatatggtga aaccccgtct
     1021 ctaccaaaaa aacaaaaatt agctgagcct ggtcatgcat gcctggaatc ccaacaactc
     1081 gggaggctga ggcaggagaa tcgcttgaac ccaggaggcg gagattgcag tgagccaaga
     1141 ttgtgccact gcactccagc ttggttccca atagaccccg caggccctac aggttgtctt
     1201 cccaacttgc cccttgctcc ataccacccc cctccacccc ataatattat agaaggacac
     1261 ctagtcagac aaaatgatgc aacttaattt tattaggaca aggctggtgg gcactggagt
     1321 ggcaacttcc agggccagga gaggcactgg ggaggggtca cagggatgcc acccgtagat
     1381 ctctcgagca gcgctcggta tcgatgcggg gaggcggccc aaagggagat ccgactcgtc
     1441 tgagggcgaa ggcgaagacg cggaagaggc cgcagagccg gcagcaggcc gcgggaagga
     1501 aggtccgctg gattgagggc cgaagggacg tagcagaagg acgtcccgcg cagaatccag
     1561 gtggcaacat aggcgagcag ccaaggaaag gacgatgatt tccccgacaa caccacggaa
     1621 ttgtcagtgc ccaacagccg agcccctgtc cagcagcggg caaggcaggc ggcgatgagt
     1681 tccgccgtgg caatagggag ggggaaagcg aaagtcccgg aaaggagctg acaggtggtg
     1741 gcaatgcccc aaccagtggg ggttgcgtca gcaaacacag tgcacaccac gccacgttgc
     1801 ctgacaacgg gccacaactc ctcataaaga gacagcaacc aggatttata caaggaggag
     1861 aaaatgaaag ccatacggga agcaatagca tgatacaaag gcattaaagc agcgtatcca
     1921 catagcgtaa aaggagcaac atagttaaga ataccagtca atctttcaca aattttgtaa
     1981 tccagaggtt gattatcgat aagcttgata tcgaattcat aacttcgtat agcatacatt
     2041 atacgaagtt attctttgca ccattctaaa gaataacagt gataatttct gggttaaggc
     2101 aaataacttc gtataggata ctttatacga agttatgcta gccttacttg tacagctcgt
     2161 ccatgccgcc ggtggagtgg cggccctcgg cgcgttcgta ctgttccacg atggtgtagt
     2221 cctcgttgtg ggaggtgatg tccaacttga tgttgacgtt gtaggcgccg ggcagctgca
     2281 cgggcttctt ggccttgtag gtggtcttga cctcagcgtc gtagtggccg ccgtccttca
     2341 gcttcagcct ctgcttgatc tcgcccttca gggcgccgtc ctcggggtac atccgctcgg
     2401 aggaggcctc ccagcccatg gtcttcttct gcattacggg gccgtcggag gggaagttgg
     2461 tgccgcgcag cttcaccttg tagatgaact cgccgtcctg cagggaggag tcctgggtca
     2521 cggtcaccac gccgccgtcc tcgaagttca tcacgcgctc ccacttgaag ccctcgggga
     2581 aggacagctt caagtagtcg gggatgtcgg cggggtgctt cacgtaggcc ttggagccgt
     2641 acatgaactg aggggacagg atgtcccagg cgaagggcag ggggccaccc ttggtcacct
     2701 tcagcttggc ggtctgggtg ccctcgtagg ggcggccctc gccctcgccc tcgatctcga
     2761 actcgtggcc gttcacggag ccctccatgt gcaccttgaa gcgcatgaac tccttgatga
     2821 tggccatgtt atcctcctcg cccttgctca ccatggtggc gaccggggga tccttcaagg
     2881 cctgctcggg tgcgcgcttg tgaaaaatga ccgactgtct ctgctggtac tgctgcttgc
     2941 gcctcttttt tttgtcacac tggcacagca gcagcatctt gaaagtggtt ctgaatgttt
     3001 tgttgcacag agcatagcac acggggttca cggtgctgtt gatgtagcac agccagtagc
     3061 ccagattcca aaaggttttg ggtatgcagc tgtcacaaaa ggtgttcacc agaaccatga
     3121 tgttgtatgg ggtccaagtg atgatgaagg caagcaagat cgcactgagg gtctgggccg
     3181 ctttcttctc cttgaccagg gacatccttt tccgcttagt gatctgactt ctggtcttca
     3241 gagcaaacct cttggccaga gtggcttcct tgaaggacag aggtagagtg gccgtgctct
     3301 tacccactga ggagttgacg tcagaagtct tagctgtgtc cacggctgac tctagctgga
     3361 tgggaagctt ggagaagctt tttggaaaac tgcctccatc gtccacgctc ttctgggcct
     3421 gcagcttgtc ggctttcctc tccaagtcca ccatccccag ctcctcctca ggcacctgca
     3481 ggttgtccga tgagggtaac ttggtggagt tgaggatggt gctgtgaccc ggaagcttga
     3541 gcacgatgga gtagatggct ctcgtctcgg agccaatgtc ctcctcgtcg gaggaggcgg
     3601 agttctccag ggaggcagca gcatcattgt tgttccaact gtcactgctg ctgtggtctt
     3661 ggtccatctg ctcggagctg ggtttccagc tcttggttgt gaaccagaag tggcagcggc
     3721 catacttcct cctgttggag cgtttcatgc tttgctgttg aagttcgtaa ctgctgcagc
     3781 ttcgagaact gcccgtgggg tggacaaagt tttctgtctc tgcctctgtc ccagaggctt
     3841 gcaggccagc aagctctttg gtacgctttt cagtttcctt atagatcctc cagtataaaa
     3901 tagtcataat ggtgacaggc atataaaaac cagcgatggc tgtgccaaaa gtaatggtgg
     3961 gctcactgag gaactgaatg aagcactctc ccggaggcac agttctcttt ccaacaaagt
     4021 attgccagaa caagatggca ggagcccaaa ggacaaagga gatgacccaa gccagaccga
     4081 tcatcacacc ggctctcttt gttgttcgtt tggctcggta cgtgagcggc ctcgtgatgg
     4141 aaaagtatct gtcaaagctg atgaccagaa gattcataac agaggcattg ctggctacgc
     4201 agtcaatggc aagccagagg tcacaggcca agttccctaa ggcccatcga ttcatgatga
     4261 tgtaggtcgt aaacagattc attgaaatga ccccgataat cagatcggca caggccaggc
     4321 ttaagaggaa gtagttgttg accgtcttca gctgcttgtt gaccttaaat gacacaatta
     4381 ccaggatgtt gccgatgatg gtcaccaagg ccaggatgcc cgttaagaaa gcgatgaaga
     4441 ccacttgcca gacggtatga cctcccagag ggtcatcggt ggtaccgtct ggagaggaga
     4501 aattgccagc tgctcgagaa acattgtagc tgccgaaatg agtgacggtt cccgggggca
     4561 gccctgcatc ggaggggctg tgtatccagg aggagctgat gtttggaaac aaaggcgagg
     4621 ttgtactgtt attgtgcaag gtcatggtgg cgcgccataa cttcgtataa tgtatgctat
     4681 acgaagttat taagaggttt catattgcta atagcagcta caatccagct accattctgc
     4741 ataacttcgt ataaagtatc ctatacgaag ttattccgga gtcgaccccc gggtaccgag
     4801 ctcgaattct ccaggcgatc tgacggttca ctaaacgagc tctgcttata taggcctccc
     4861 accgtacacg cctacctcga catacgttct ctatcactga tagggagtaa actcgacata
     4921 cgttctctat cactgatagg gataaactcg acatacgttc tctatcactg atagggagta
     4981 aactcgacat acgttctcta tcactgatag ggagtaaact cgacatacgt tctctatcac
     5041 tgatagggag taaactcgac atcgttctct atcactgata gggagtaaac tcgacatacg
     5101 ttctctatca ctgataggga gtaaacgcgt gcggccgcag gaacccctag tgatggagtt
     5161 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg
     5221 acgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcaggac atgtgagcaa
     5281 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     5341 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     5401 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5461 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5521 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5581 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5641 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5701 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5761 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5821 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5881 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5941 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     6001 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     6061 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     6121 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     6181 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     6241 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     6301 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6361 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6421 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6481 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6541 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6601 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6661 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6721 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6781 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6841 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6901 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6961 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     7021 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     7081 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     7141 cctttcgt
//
