LOCUS       sequence-394293-        4822 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of mAmetrine1.1 under the TRE promoter in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4822)
  AUTHORS   Leiwe MN, Fujimoto S, Baba T, Moriyasu D, Saha B, Sakaguchi R, 
            Inagaki S, Imai T
  TITLE     Automated neuronal reconstruction with super-multicolour Tetbow 
            labelling and threshold-based clustering of colour hues.
  JOURNAL   Nat Commun. 2024 Jun 25;15(1):5279. doi: 10.1038/s41467-024-49455-y.
  PUBMED    38918382
REFERENCE   2  (bases 1 to 4822)
  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: "Nat 
            Commun."; date: "2024-06-25"; volume: "15(1):5279. doi"; pages: " 
            10.1038/s41467-024-49455-y"
COMMENT     Sequence Label: pAAV-TRE-mAmetrine1.1-WPRE
FEATURES             Location/Qualifiers
     source          1..4822
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   177..306
                     /label=AAV2 ITR
     polyA_signal    468..589
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(505..524)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     559..578
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     618..634
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     619..635
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    638..1226
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             744..755
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1153..1173
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(1240..1953,1954..1956,1957..1959))
                     /codon_start=1
                     /product="photostable yellow fluorescent protein with a 
                     large Stokes shift, excitable with violet light (Ai et al.,
                     2008)"
                     /label=mAmetrine1.1
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVRGEGEGDATNGKLTL
                     KFICTSGELPVPWPTLVTTLSYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GSYRTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNMNAWDAYITADKQKNGIK
                     ANFKIEHNVKDGGVQLADAYQQNTPIGDGSVLLPDNHYLSFQSKLFKDPNEQRDHMVLL
                     EFVTAAGITPGMDELYK"
     primer_bind     1893..1914
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1956..1965
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1979..2003)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2000..2038)
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    2050..2068
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2086..2104
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2121..2139
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2157..2175
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2193..2211
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2228..2246
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2264..2282
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     repeat_region   2356..2485
                     /label=AAV2 ITR
     rep_origin      2557..3012
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2644..2663)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2854..2875
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3038..3142
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3143..3211,3212..4003)
                     /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(3361..3380)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4174..4762
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4663..4682
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
       61 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      121 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgc
      181 gcgctcgctc gctcactgag gccgcccggg caaagcccgg gcgtcgggcg acctttggtc
      241 gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc atcactaggg
      301 gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca tgctctagga
      361 agatccttat cgattttacc acatttgtag aggttttact tgctttaaaa aacctcccac
      421 atctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg
      481 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
      541 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgct
      601 cgaagcggcc gcccgggtcg aggtcgacgg tatcgatgcg gggaggcggc ccaaagggag
      661 atccgactcg tctgagggcg aaggcgaaga cgcggaagag gccgcagagc cggcagcagg
      721 ccgcgggaag gaaggtccgc tggattgagg gccgaaggga cgtagcagaa ggacgtcccg
      781 cgcagaatcc aggtggcaac acaggcgagc agccaaggaa aggacgatga tttccccgac
      841 aacaccacgg aattgtcagt gcccaacagc cgagcccctg tccagcagcg ggcaaggcag
      901 gcggcgatga gttccgccgt ggcaataggg agggggaaag cgaaagtccc ggaaaggagc
      961 tgacaggtgg tggcaatgcc ccaaccagtg ggggttgcgt cagcaaacac agtgcacacc
     1021 acgccacgtt gcctgacaac gggccacaac tcctcataaa gagacagcaa ccaggattta
     1081 tacaaggagg agaaaatgaa agccatacgg gaagcaatag catgatacaa aggcattaaa
     1141 gcagcgtatc cacatagcgt aaaaggagca acatagttaa gaataccagt caatctttca
     1201 caaattttgt aatccagagg ttgattatcg ataagctttt cacttgtaca gctcgtccat
     1261 gccgggagtt atcccggcgg cggtaacgaa ctccagcagg accatgtgat cgcgctgctc
     1321 gttggggtct ttgaacagct tgctctggaa gctcaggtag tggttgtcag gcagcagcac
     1381 ggagccgtcg ccgatggggg tgttctgctg gtacgcgtcg gcgagctgca cgccgccgtc
     1441 cttgacgttg tgctcgatct tgaagttcgc tttgatgccg ttcttctgct tgtcggccgt
     1501 gatatacgcg tcccacgcgt tcatgttgta ctccagcttg tgccccagga tgttgccgtc
     1561 ctccttgaag tcgatgccct tcagctcgat gcggttcacc agggtgtcgc cctcgaactt
     1621 cacctcggcg cgggtcctgt agctgccgtc gtccttgaag gagatggtgc gctcctggac
     1681 gtagccttcg ggcatggcgg acttgaagaa gtcgtgctgc ttcatgtggt cggggtagcg
     1741 ggcgaagcac tgcacgccgt aagacagggt ggtcacgagg gtgggccagg gcacgggcag
     1801 ctcgccggag gtgcagatga acttcagggt cagcttgccg ttggtggcat cgccctcgcc
     1861 ctcgccgcgc acgctgaacc tgtggccgtt tacgtcgccg tccagctcga ccaggatggg
     1921 caccaccccg gtgaacagct cctcgccctt gctcaccatg gtggcgaatt ctccaggcga
     1981 tctgacggtt cactaaacga gctctgctta tataggcctc ccaccgtaca cgcctacctc
     2041 gacatacgtt ctctatcact gatagggagt aaactcgaca tacgttctct atcactgata
     2101 gggataaact cgacatacgt tctctatcac tgatagggag taaactcgac atacgttctc
     2161 tatcactgat agggagtaaa ctcgacatac gttctctatc actgataggg agtaaactcg
     2221 acatcgttct ctatcactga tagggagtaa actcgacata cgttctctat cactgatagg
     2281 gagtaaacca attcgagatc ttcctagagc atggctacgt agataagtag catggcgggt
     2341 taatcattaa ctacaaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc
     2401 gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg
     2461 cctcagtgag cgagcgagcg cgcagctggc gtaatagcga agaggcccgc accgatcgcc
     2521 cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa
     2581 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     2641 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     2701 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     2761 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     2821 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     2881 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     2941 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     3001 cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     3061 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     3121 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     3181 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     3241 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     3301 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     3361 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     3421 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     3481 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     3541 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     3601 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     3661 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     3721 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     3781 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     3841 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     3901 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     3961 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     4021 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     4081 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     4141 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     4201 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     4261 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     4321 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     4381 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     4441 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     4501 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     4561 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     4621 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     4681 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     4741 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     4801 ttgctggcct tttgctcaca tg
//
