LOCUS       sequence-365752-        6284 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express GCaMP6f under TetO(3G).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6284)
  AUTHORS   Amo R, Matias S, Yamanaka A, Tanaka KF, Uchida N, Watabe-Uchida M
  TITLE     A gradual temporal shift of dopamine responses mirrors the 
            progression of temporal difference error in machine learning.
  JOURNAL   Nat Neurosci. 2022 Aug;25(8):1082-1092. doi: 
            10.1038/s41593-022-01109-2. Epub 2022 Jul 7.
  PUBMED    35798979
REFERENCE   2  (bases 1 to 6284)
  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/s41593-022-01109-2"; journalName: "Nat Neurosci"; date: 
            "2022-08"; volume: "25"; issue: "8"; pages: "1082-1092"
COMMENT     Sequence Label: pAAV-TetO(3G)-GCaMP6f-WPRE
FEATURES             Location/Qualifiers
     source          1..6284
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   5..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     protein_bind    150..168
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    186..204
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    222..240
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    258..276
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    294..312
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    330..348
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    366..384
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     430..454
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1066..1075
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1072..1080,1081..1098,1099..1101,1102..1134,1135..1137,
                     1138..1161,1162..1188,1189..1245,1246..1251,1252..1521,
                     1522..1539,1540..1974,1975..1980,1981..2424)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      1168..1177
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1458..1479
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1585..1606)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1846..1865)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    2446..3034
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2499..2519)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2917..2928)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3046..3522
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3245..3264)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     rep_origin      3766..4221
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3853..3872)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4063..4084
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4238..4257)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4357..4379
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4417..4435)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4503..4607
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4608..4676,4677..5468)
                     /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(4826..4845)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5639..6227
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6128..6147
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgtcctgca ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc gtcgggcgac
       61 ctttggtcgc ccggcctcag tgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgttttact ccctatcagt gatagagaac gtatgaagag
      181 tttactccct atcagtgata gagaacgtat gcagacttta ctccctatca gtgatagaga
      241 acgtataagg agtttactcc ctatcagtga tagagaacgt atgaccagtt tactccctat
      301 cagtgataga gaacgtatct acagtttact ccctatcagt gatagagaac gtatatccag
      361 tttactccct atcagtgata gagaacgtat gtcgaggtag gcgtgtacgg tgggcgccta
      421 taaaagcaga gctcgtttag tgaaccgtca gatcgcctgg agcaattcca caacactttt
      481 gtcttatacc aactttccgt accacttcct accctcgttg agaacttcag ggtgagtttg
      541 gggacccttg attgttcttt ctttttcgct attgtaaaat tcatgttata tggagggggc
      601 aaagttttca gggtgttgtt tagaatggga agatgtccct tgtatcacca tggaccctca
      661 tgataatttt gtttctttca ctttctactc tgttgacaac cattgtctcc tcttattttc
      721 ttttcatttt ctgtaacttt ttcgttaaac tttagcttgc atttgtaacg aatttttaaa
      781 ttcacttttg tttatttgtc agattgtaag tactttctct aatcactttt ttttcaaggc
      841 aatcagggta tattatattg tacttcagca cagttttaga gaacaattgt tataattaaa
      901 tgataaggta gaatatttct gcatataaat tctggctggc gtggaaatat tcttattggt
      961 agaaacaact acatcctggt catcatcctg cctttctcct tatggttaca atgatataca
     1021 ctgtttgaga tgaggataaa atactctgag tccaaaccgg atctcgccac catgggttct
     1081 catcatcatc atcatcatgg tatggctagc atgactggtg gacagcaaat gggtcgggat
     1141 ctgtacgacg atgacgataa ggatctcgcc accatggtcg actcatcacg tcgtaagtgg
     1201 aataagacag gtcacgcagt cagagctata ggtcggctga gctcactcga gaacgtctat
     1261 atcaaggccg acaagcagaa gaacggcatc aaggcgaact tcaagatccg ccacaacatc
     1321 gaggacggcg gcgtgcagct cgcctaccac taccagcaga acacccccat cggcgacggc
     1381 cccgtgctgc tgcccgacaa ccactacctg agcgtgcagt ccaaactttc gaaagacccc
     1441 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     1501 ggcatggacg agctgtacaa gggcggtacc ggagggagca tggtgagcaa gggcgaggag
     1561 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1621 ttcagcgtgt ccggcgaggg tgagggcgat gccacctacg gcaagctgac cctgaagttc
     1681 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     1741 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1801 gccatgcccg aaggctacat ccaggagcgc accatcttct tcaaggacga cggcaactac
     1861 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1921 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caacctgccg
     1981 gaccaactga ctgaagagca gatcgcagaa tttaaagagg aattctccct atttgacaag
     2041 gacggggatg ggacaataac aaccaaggag ctggggacgg tgatgcggtc tctggggcag
     2101 aaccccacag aagcagagct gcaggacatg atcaatgaag tagatgccga cggtgacggc
     2161 acaatcgact tccctgagtt cctgacaatg atggcaagaa aaatgaaata cagggacacg
     2221 gaagaagaaa ttagagaagc gttcggtgtg tttgataagg atggcaatgg ctacatcagt
     2281 gcagcagagc ttcgccacgt gatgacaaac cttggagaga agttaacaga tgaagaggtt
     2341 gatgaaatga tcagggaagc agacatcgat ggggatggtc aggtaaacta cgaagagttt
     2401 gtacaaatga tgacagcgaa gtgagcggcc tcgactgaat tcgataatca acctctggat
     2461 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt
     2521 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc
     2581 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg
     2641 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc
     2701 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa
     2761 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat
     2821 tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgccta tgttgccacc
     2881 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt
     2941 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag
     3001 acgagtcgga tctccctttg ggccgcctcc ccgcatcgga tctacgggtg gcatccctgt
     3061 gacccctccc cagtgcctct cctggccctg gaagttgcca ctccagtgcc caccagcctt
     3121 gtcctaataa aattaagttg catcattttg tctgactagg tgtccttcta taatattatg
     3181 gggtggaggg gggtggtatg gagcaagggg caagttggga agacaacctg tagggcctgc
     3241 ggggtctatt gggaaccaag ctggagtgca gtggcacaat cttggctcac tgcaatctcc
     3301 gcctcctggg ttcaagcgat tctcctgcct cagcctcccg agttgttggg attccaggca
     3361 tgcatgacca ggctcagcta atttttgttt ttttggtaga gacggggttt caccatattg
     3421 gccaggctgg tctccaactc ctaatctcag gtgatctacc caccttggcc tcccaaattg
     3481 ctgggattac aggcgtgaac cactgctccc ttccctgtcc ttctgatttt gtaggtaacc
     3541 acgtgcggac cgagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc
     3601 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggcggcctca
     3661 gtgagcgagc gagmgnrsas mkgcctgcag gggcgcctga tgcggtattt tctccttacg
     3721 catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg
     3781 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     3841 ccttagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     3901 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     3961 tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga
     4021 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     4081 ctggaacaac actcaactct atctcgggct attcttttga tttataaggg attttgccga
     4141 tttcggtcta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     4201 aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat
     4261 agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     4321 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     4381 tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat
     4441 aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg
     4501 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     4561 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     4621 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     4681 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     4741 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     4801 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     4861 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     4921 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     4981 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     5041 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     5101 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     5161 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     5221 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     5281 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     5341 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     5401 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     5461 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     5521 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     5581 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     5641 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     5701 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     5761 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     5821 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     5881 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     5941 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     6001 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     6061 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     6121 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     6181 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     6241 cggccttttt acggttcctg gccttttgct ggccttttgc tcac
//
