LOCUS       sequence-394048-        4752 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses 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 4752)
  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 4752)
  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: pBS-TRE-mAmetrine1.1-WPRE
FEATURES             Location/Qualifiers
     source          1..4752
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    659..677
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    695..713
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    730..748
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    766..784
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    802..820
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    837..855
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    873..891
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        903..941
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     938..962
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      976..985
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(982..984,985..987,988..1701)
                     /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     complement(1027..1048)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     misc_feature    1720..2308
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1773..1793)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2191..2202)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2469..2550
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2506..2525)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2564..2584)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2564..2582)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2603..2619)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2603..2619)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2616..2638)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2627..2643
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2651..2657,2658..2675,2676..2681))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2696..2717
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2834..2851)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3005..3593)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3085..3104)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3764..4555,4556..4624))
                     /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     4387..4406
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4625..4729)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggtttactc
      661 cctatcagtg atagagaacg tatgtcgagt ttactcccta tcagtgatag agaacgatgt
      721 cgagtttact ccctatcagt gatagagaac gtatgtcgag tttactccct atcagtgata
      781 gagaacgtat gtcgagttta ctccctatca gtgatagaga acgtatgtcg agtttatccc
      841 tatcagtgat agagaacgta tgtcgagttt actccctatc agtgatagag aacgtatgtc
      901 gaggtaggcg tgtacggtgg gaggcctata taagcagagc tcgtttagtg aaccgtcaga
      961 tcgcctggag aattcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg
     1021 cccatcctgg tcgagctgga cggcgacgta aacggccaca ggttcagcgt gcgcggcgag
     1081 ggcgagggcg atgccaccaa cggcaagctg accctgaagt tcatctgcac ctccggcgag
     1141 ctgcccgtgc cctggcccac cctcgtgacc accctgtctt acggcgtgca gtgcttcgcc
     1201 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     1261 gtccaggagc gcaccatctc cttcaaggac gacggcagct acaggacccg cgccgaggtg
     1321 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     1381 gacggcaaca tcctggggca caagctggag tacaacatga acgcgtggga cgcgtatatc
     1441 acggccgaca agcagaagaa cggcatcaaa gcgaacttca agatcgagca caacgtcaag
     1501 gacggcggcg tgcagctcgc cgacgcgtac cagcagaaca cccccatcgg cgacggctcc
     1561 gtgctgctgc ctgacaacca ctacctgagc ttccagagca agctgttcaa agaccccaac
     1621 gagcagcgcg atcacatggt cctgctggag ttcgttaccg ccgccgggat aactcccggc
     1681 atggacgagc tgtacaagtg agcggccgcg cttatcgata atcaacctct ggattacaaa
     1741 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     1801 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc
     1861 ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt
     1921 ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc
     1981 tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc
     2041 gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg
     2101 gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc cacctggatt
     2161 ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc
     2221 cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt
     2281 cggatctccc tttgggccgc ctccccgcat cgataccgtc gacgcggccg catcgataag
     2341 cttgtcgacg atatctctag aggatcataa tcagccatac cacatttgta gaggttttac
     2401 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     2461 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     2521 atttcacaaa taaagcattt ttttcactgc ccagcttttg ttccctttag tgagggttaa
     2581 ttgcgcgctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     2641 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     2701 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     2761 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     2821 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     2881 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     2941 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     3001 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     3061 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     3121 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     3181 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     3241 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     3301 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     3361 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     3421 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     3481 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     3541 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     3601 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     3661 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     3721 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     3781 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     3841 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     3901 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     3961 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     4021 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     4081 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     4141 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     4201 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     4261 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     4321 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     4381 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     4441 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     4501 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     4561 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     4621 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     4681 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     4741 gaaaagtgcc ac
//
