LOCUS       sequence-416392-        6337 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  control for targeted actin nucleation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6337)
  AUTHORS   Nakamura H, Rho E, Lee CT, Itoh K, Deng D, Watanabe S, Razavi S, 
            Matsubayashi HT, Zhu C, Jung E, Rangamani P, Watanabe S, Inoue T
  TITLE     ActuAtor, a Listeria-inspired molecular tool for physical 
            manipulation of intracellular organizations through de novo actin 
            polymerization.
  JOURNAL   Cell Rep. 2023 Sep 2:113089. doi: 10.1016/j.celrep.2023.113089.
  PUBMED    37734382
REFERENCE   2  (bases 1 to 6337)
  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: "Cell Rep. 
            2023 Sep 2:113089. doi: 10.1016/j.celrep.2023.113089."
COMMENT     Sequence Label: EYFP-FKBP-DN-N-WASP
FEATURES             Location/Qualifiers
     source          1..6337
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      3..12
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(51..254)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(80..100)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        255..558
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(733..1321)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(813..832)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    1650..1697
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     1722..1741
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(1929..2723)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(1929..2723)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2040..2059)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2650..2669
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(2758..3115)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2772..2907
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        2924..3115
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     3072..3092
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(3117..3221)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3248..3703
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3335..3354)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3545..3566
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    3710..3831
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3721..3740)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3775..3794
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5294..5614)
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             complement(join(5627..6337,1..3,4..6))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5669..5690)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     6018..6037
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6277..6298
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
ORIGIN
        1 caccatggtg gcgaccggta gcgctagcgg atctgacggt tcactaaacc agctctgctt
       61 atatagacct cccaccgtac acgcctaccg cccatttgcg tcaatggggc ggagttgtta
      121 cgacattttg gaaagtcccg ttgattttgg tgccaaaaca aactcccatt gacgtcaatg
      181 gggtggagac ttggaaatcc ccgtgagtca aaccgctatc cacgcccatt gatgtactgc
      241 caaaaccgca tcaccatggt aatagcgatg actaatacgt agatgtactg ccaagtagga
      301 aagtcccata aggtcatgta ctgggcataa tgccaggcgg gccatttacc gtcattgacg
      361 tcaatagggg gcgtacttgg catatgatac acttgatgta ctgccaagtg ggcagtttac
      421 cgtaaatact ccacccattg acgtcaatgg aaagtcccta ttggcgttac tatgggaaca
      481 tacgtcatta ttgacgtcaa tgggcggggg tcgttgggcg gtcagccagg cgggccattt
      541 accgtaagtt atgtaacgcg gaactccata tatgggctat gaactaatga ccccgtaatt
      601 gattactatt aataactaat gcatggcggt aatacggtta tccacagaat caggggataa
      661 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
      721 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
      781 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
      841 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
      901 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
      961 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     1021 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     1081 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     1141 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     1201 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     1261 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     1321 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     1381 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     1441 atgaagtttt aaatcaatct aaagtatata tgagtaacct gaggctatgg cagggcctgc
     1501 cgccccgacg ttggctgcga gccctgggcc ttcacccgaa cttggggggt ggggtgggga
     1561 aaaggaagaa acgcgggcgt attggcccca atggggtctc ggtggggtat cgacagagtg
     1621 ccagccctgg gaccgaaccc cgcgtttatg aacaaacgac ccaacaccgt gcgttttatt
     1681 ctgtcttttt attgccgtca tagcgcgggt tccttccggt attgtctcct tccgtgtttc
     1741 agttagcctc cccctagggt gggcgaagaa ctccagcatg agatccccgc gctggaggat
     1801 catccagccg gcgtcccgga aaacgattcc gaagcccaac ctttcataga aggcggcggt
     1861 ggaatcgaaa tctcgtgatg gcaggttggg cgtcgcttgg tcggtcattt cgaaccccag
     1921 agtcccgctc agaagaactc gtcaagaagg cgatagaagg cgatgcgctg cgaatcggga
     1981 gcggcgatac cgtaaagcac gaggaagcgg tcagcccatt cgccgccaag ctcttcagca
     2041 atatcacggg tagccaacgc tatgtcctga tagcggtccg ccacacccag ccggccacag
     2101 tcgatgaatc cagaaaagcg gccattttcc accatgatat tcggcaagca ggcatcgcca
     2161 tgggtcacga cgagatcctc gccgtcgggc atgctcgcct tgagcctggc gaacagttcg
     2221 gctggcgcga gcccctgatg ctcttcgtcc agatcatcct gatcgacaag accggcttcc
     2281 atccgagtac gtgctcgctc gatgcgatgt ttcgcttggt ggtcgaatgg gcaggtagcc
     2341 ggatcaagcg tatgcagccg ccgcattgca tcagccatga tggatacttt ctcggcagga
     2401 gcaaggtgag atgacaggag atcctgcccc ggcacttcgc ccaatagcag ccagtccctt
     2461 cccgcttcag tgacaacgtc gagcacagct gcgcaaggaa cgcccgtcgt ggccagccac
     2521 gatagccgcg ctgcctcgtc ttgcagttca ttcagggcac cggacaggtc ggtcttgaca
     2581 aaaagaaccg ggcgcccctg cgctgacagc cggaacacgg cggcatcaga gcagccgatt
     2641 gtctgttgtg cccagtcata gccgaatagc ctctccaccc aagcggccgg agaacctgcg
     2701 tgcaatccat cttgttcaat catgcgaaac gatcctcatc ctgtctcttg atcgatcttt
     2761 gcaaaagcct aggcctccaa aaaagcctcc tcactacttc tggaatagct cagaggccga
     2821 ggcggcctcg gcctctgcat aaataaaaaa aattagtcag ccatggggcg gagaatgggc
     2881 ggaactgggc ggagttaggg gcgggatggg cggagttagg ggcgggacta tggttgctga
     2941 ctaattgaga tgcatgcttt gcatacttct gcctgctggg gagcctgggg actttccaca
     3001 cctggttgct gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg
     3061 ggactttcca caccctaact gacacacatt ccacagctgg ttctttccgc ctcaggactc
     3121 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     3181 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     3241 ccacctgacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     3301 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     3361 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     3421 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     3481 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     3541 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     3601 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     3661 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttacgcctt aagatacatt
     3721 gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt
     3781 tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac
     3841 aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag
     3901 taaaacctct acaaatgtgg tatggctgat tatgatcagt tatctagatc cggtggatcc
     3961 cgggcccgcg gtaccgtcga ctgcagaatt ctcagtcttc ccattcatca tcatcctcaa
     4021 aatcttcatc atcatcttca tcctcatctt cgtctgaaga atgaatggct ttgctcctct
     4081 tctgcatcac ttccattaat gcacctacaa ttcctgaagt gggtgcaggt gctggtggtg
     4141 tagactctgg ggcatcagtt acagatttca gttgaatacc ctgtcgtatc tggtctaaaa
     4201 gtgcatccct tccagagcag gacaccggcc gactgttctg ttccactttt tttagctgag
     4261 caccctctct aatttgatct aaaagagctg ctttgcttcc tgcaggagtt ggaacttggt
     4321 ggtcaccatc agaagggagg ccagggggag gtggtggccc tggtggaggt ggaggtggcg
     4381 gcggaggtgg tggtgccact gacccgctca ctgacagagg tggaggtggt ggtggaggcc
     4441 ctgaaggtgc tgaggaggga agagctggaa gtggaggagg gtacatcctg tttggcggtg
     4501 gtggaggggc tcctacacct ggcctggatg gaggcggtgg tggcggtgca gctgtgggag
     4561 ctcttgaagg tggtggagga ggagcccctc ttcccctggc aggaggggga ggagggcccg
     4621 agctatgtgg aggcggggga ggagggggcg gccctcccct tgatggtggt ggaggtggtg
     4681 gtgcttgcct tcgcagttca tttttaacag cttcaacacc tcctgttttt tcaatgaagt
     4741 catatataac ttttgatgtt tctctgtctt taagttgtgc ctctgagatt ccacacatat
     4801 caaaaagatt cttcaattct ggatccaaat tattcagatc aaagccagta ttcggatccc
     4861 aacccacatg tccaatgtgt tggaaattgc ttggtgttcc aatatctgcc ttagttaacc
     4921 tcttcttttt agcttttcct tttttctttt ctttggtatg ggagatgttg ttgatttgcg
     4981 gaccataaaa tctatttgtt gtaatttctg gattttttat gtcaactgtt gccatgggta
     5041 gattaggacc atttgggggg tctcgtcttt tctcagattt cctttgtcgt cgtcccaaca
     5101 agtctgtaac tgcttttcgg aatttttttg cttcttcttc attggcaaaa ttaagagcaa
     5161 cttgacaggt atctccagca aaggtatgaa aatatcctct aggactatta tatacggaag
     5221 cttgagctcg aggaccacca gcactaccac cagcactacc accagcacta ccaccagcac
     5281 taccaccagc actttccagt tttagaagct ccacatcgaa gacgagagtg gcatgtggtg
     5341 ggatgatgcc tgggtgccca gtggcaccat aggcataatc tggagatata gtcagtttgg
     5401 ctctctgacc cacactcatc tgggcaaccc cttcttccca gcctcggatc acctcctgct
     5461 tgcctagcat aaacttaaag ggcttgtttc tgtcccggga ggaatcaaat ttctttccat
     5521 cttcaagcat cccggtgtag tgcaccacgc aggtctggcc gcgcttgggg aaggtgcgcc
     5581 cgtctcctgg ggagatggtt tccacctgca ctccaccacc agcactcttg tacagctcgt
     5641 ccatgccgag agtgatcccg gcggcggtca cgaactccag caggaccatg tgatcgcgct
     5701 tctcgttggg gtctttgctc agggcggact ggtagctcag gtagtggttg tcgggcagca
     5761 gcacggggcc gtcgccgatg ggggtgttct gctggtagtg gtcggcgagc tgcacgctgc
     5821 cgtcctcgat gttgtggcgg atcttgaagt tcaccttgat gccgttcttc tgcttgtcgg
     5881 ccatgatata gacgttgtgg ctgttgtagt tgtactccag cttgtgcccc aggatgttgc
     5941 cgtcctcctt gaagtcgatg cccttcagct cgatgcggtt caccagggtg tcgccctcga
     6001 acttcacctc ggcgcgggtc ttgtagttgc cgtcgtcctt gaagaagatg gtgcgctcct
     6061 ggacgtagcc ttcgggcatg gcggacttga agaagtcgtg ctgcttcatg tggtcggggt
     6121 agcgggcgaa gcactgcagg ccgtagccga aggtggtcac gagggtgggc cagggcacgg
     6181 gcagcttgcc ggtggtgcag atgaacttca gggtcagctt gccgtaggtg gcatcgccct
     6241 cgccctcgcc ggacacgctg aacttgtggc cgtttacgtc gccgtccagc tcgaccagga
     6301 tgggcaccac cccggtgaac agctcctcgc ccttgct
//
