LOCUS       sequence-394416-        5620 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of tdKatushka2 under the TRE promoter in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5620)
  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 5620)
  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-tdKatushka2-WPRE
FEATURES             Location/Qualifiers
     source          1..5620
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   51..180
                     /label=AAV2 ITR
     protein_bind    254..272
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    290..308
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    325..343
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    361..379
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    397..415
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    432..450
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    468..486
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        498..536
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     533..557
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     complement(584..600)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(584..600)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             621..1325
                     /codon_start=1
                     /product="improved version of the oligomeric far-red 
                     fluorescent protein TurboFP635/Katushka (Luker et al., 
                     2015)"
                     /label=Katushka2S
                     /note="mammalian codon-optimized"
                     /translation="MVGEDSVLITENMHMKLYMEGTVNDHHFKCTSEGEGKPYEGTQTM
                     KIKVVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERITTYEDGG
                     VLTATQDTSLQNGCLIYNVKINGVNFPSNGPVMQKKTLGWEASTEMLYPADSGLRGHAQ
                     MALKLVGGGYLHCSLKTTYRSKKPAKNLKMPGFYFVDRRLERIKEADKETYVEQHEMAV
                     ARYCDLPSKLGHS"
     CDS             621..1325
                     /codon_start=1
                     /product="far-red-shifted derivative of red fluorescent 
                     protein from Entacmaea quadricolor, also known as Katushka 
                     (Shcherbo et al., 2007)"
                     /label=TurboFP635
                     /note="mammalian codon-optimized"
                     /translation="MVGEDSVLITENMHMKLYMEGTVNDHHFKCTSEGEGKPYEGTQTM
                     KIKVVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERITTYEDGG
                     VLTATQDTSLQNGCLIYNVKINGVNFPSNGPVMQKKTLGWEASTEMLYPADSGLRGHAQ
                     MALKLVGGGYLHCSLKTTYRSKKPAKNLKMPGFYFVDRRLERIKEADKETYVEQHEMAV
                     ARYCDLPSKLGHS"
     CDS             1395..2099
                     /codon_start=1
                     /product="improved version of the oligomeric far-red 
                     fluorescent protein TurboFP635/Katushka (Luker et al., 
                     2015)"
                     /label=Katushka2S
                     /note="mammalian codon-optimized"
                     /translation="MVGEDSVLITENMHMKLYMEGTVNDHHFKCTSEGEGKPYEGTQTM
                     KIKVVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERITTYEDGG
                     VLTATQDTSLQNGCLIYNVKINGVNFPSNGPVMQKKTLGWEASTEMLYPADSGLRGHAQ
                     MALKLVGGGYLHCSLKTTYRSKKPAKNLKMPGFYFVDRRLERIKEADKETYVEQHEMAV
                     ARYCDLPSKLGHS"
     CDS             1395..2099
                     /codon_start=1
                     /product="far-red-shifted derivative of red fluorescent 
                     protein from Entacmaea quadricolor, also known as Katushka 
                     (Shcherbo et al., 2007)"
                     /label=TurboFP635
                     /note="mammalian codon-optimized"
                     /translation="MVGEDSVLITENMHMKLYMEGTVNDHHFKCTSEGEGKPYEGTQTM
                     KIKVVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERITTYEDGG
                     VLTATQDTSLQNGCLIYNVKINGVNFPSNGPVMQKKTLGWEASTEMLYPADSGLRGHAQ
                     MALKLVGGGYLHCSLKTTYRSKKPAKNLKMPGFYFVDRRLERIKEADKETYVEQHEMAV
                     ARYCDLPSKLGHS"
     misc_feature    2130..2718
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2183..2203)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2601..2612)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2721..2737)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2722..2738)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    2767..2888
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2778..2797)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2832..2851
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3394..3982)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3474..3493)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4153..4944,4945..5013))
                     /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     4776..4795
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5014..5118)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5144..5599)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5281..5302)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5493..5512
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctgcgcgctc
       61 gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt ggtcgcccgg
      121 cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact aggggttcct
      181 tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct aggaagatct
      241 cgaattggtt tactccctat cagtgataga gaacgtatgt cgagtttact ccctatcagt
      301 gatagagaac gatgtcgagt ttactcccta tcagtgatag agaacgtatg tcgagtttac
      361 tccctatcag tgatagagaa cgtatgtcga gtttactccc tatcagtgat agagaacgta
      421 tgtcgagttt atccctatca gtgatagaga acgtatgtcg agtttactcc ctatcagtga
      481 tagagaacgt atgtcgaggt aggcgtgtac ggtgggaggc ctatataagc agagctcgtt
      541 tagtgaaccg tcagatcgcc tggagaattc ctgcagcccg ggggatccac tagttctaga
      601 tttaagaagg agatatacat atggtgggtg aggatagcgt gctgatcacc gagaacatgc
      661 acatgaaact gtacatggag ggcaccgtga acgaccacca cttcaagtgc acatccgagg
      721 gcgaaggcaa gccctacgag ggcacccaga ccatgaagat caaggtggtc gagggcggcc
      781 ctctcccctt cgccttcgac atcctggcta ccagcttcat gtacggcagc aaaaccttta
      841 tcaaccacac ccagggcatc cccgacttct ttaagcagtc cttccctgag ggcttcacat
      901 gggagaggat caccacatac gaagacgggg gcgtgctgac cgctacccag gacaccagcc
      961 tccagaacgg ctgcctcatc tacaacgtca agatcaacgg ggtgaacttc ccatccaacg
     1021 gccctgtgat gcagaagaaa acactcggct gggaggccag caccgagatg ctgtaccccg
     1081 ctgacagcgg cctgagaggc catgcccaga tggccctgaa gctcgtgggc gggggctacc
     1141 tgcactgctc cctcaagacc acatacagat ccaagaaacc cgctaagaac ctcaagatgc
     1201 ccggcttcta cttcgtggac aggagactgg aaagaatcaa ggaggccgac aaagagacct
     1261 acgtcgagca gcacgagatg gctgtggcca ggtactgcga cctgcctagc aaactggggc
     1321 acagcgggca tggcaccggc agcaccggca gcggcagctc cggtaccgcc tcctccgagg
     1381 acaacaacat ggccatggtg ggtgaggata gcgtgctgat caccgagaac atgcacatga
     1441 aactgtacat ggagggcacc gtgaacgacc accacttcaa gtgcacatcc gagggcgaag
     1501 gcaagcccta cgagggcacc cagaccatga agatcaaggt ggtcgagggc ggccctctcc
     1561 ccttcgcctt cgacatcctg gctaccagct tcatgtacgg cagcaaaacc tttatcaacc
     1621 acacccaggg catccccgac ttctttaagc agtccttccc tgagggcttc acatgggaga
     1681 ggatcaccac atacgaagac gggggcgtgc tgaccgctac ccaggacacc agcctccaga
     1741 acggctgcct catctacaac gtcaagatca acggggtgaa cttcccatcc aacggccctg
     1801 tgatgcagaa gaaaacactc ggctgggagg ccagcaccga gatgctgtac cccgctgaca
     1861 gcggcctgag aggccatgcc cagatggccc tgaagctcgt gggcgggggc tacctgcact
     1921 gctccctcaa gaccacatac agatccaaga aacccgctaa gaacctcaag atgcccggct
     1981 tctacttcgt ggacaggaga ctggaaagaa tcaaggaggc cgacaaagag acctacgtcg
     2041 agcagcacga gatggctgtg gccaggtact gcgacctgcc tagcaaactg gggcacagct
     2101 ccggactcag atcttgaaag cttatcgata atcaacctct ggattacaaa atttgtgaaa
     2161 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa
     2221 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat
     2281 cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt
     2341 gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc
     2401 tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc
     2461 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg
     2521 ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc cacctggatt ctgcgcggga
     2581 cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc
     2641 tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc
     2701 tttgggccgc ctccccgcat cgataccgtc gacctcgacc cgggcggccg cttcgagcag
     2761 acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat
     2821 gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata
     2881 aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg gagatgtggg
     2941 aggtttttta aagcaagtaa aacctctaca aatgtggtaa aatcgataag gatcttccta
     3001 gagcatggct acgtagataa gtagcatggc gggttaatca ttaactacaa ggaaccccta
     3061 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgcccgggct
     3121 ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cattaatgaa tcggccaacg
     3181 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     3241 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     3301 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     3361 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     3421 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     3481 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     3541 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     3601 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     3661 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3721 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3781 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     3841 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3901 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3961 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     4021 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     4081 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     4141 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     4201 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     4261 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     4321 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     4381 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     4441 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     4501 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     4561 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     4621 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     4681 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     4741 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     4801 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     4861 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     4921 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     4981 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     5041 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     5101 acaaataggg gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat
     5161 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc
     5221 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt
     5281 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa
     5341 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg
     5401 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
     5461 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
     5521 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
     5581 gcgccgctac agggcgcgtc ccattcgcca ttcaggctgc
//
