LOCUS       sequence-481712-        6524 bp    DNA     circular SYN 01-AUG-2025
DEFINITION  Tagged vimentin for rapalog induced pulling experiments..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6524)
  AUTHORS   Pasolli M, Meiring JCM, Conboy JP, Koenderink GH, Akhmanova A
  TITLE     Optogenetic and chemical genetic tools for rapid repositioning of 
            vimentin intermediate filaments.
  JOURNAL   J Cell Biol. 2025 Sep 1;224(9):e202504004. doi: 
            10.1083/jcb.202504004. Epub 2025 Jul 8.
  PUBMED    40627079
REFERENCE   2  (bases 1 to 6524)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 1, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol."; date: "2025-09-1"; volume: "224(9):e202504004. doi"; pages: 
            " 10.1083/jcb.202504004. Epub 2025 Jul 8"
COMMENT     Sequence Label: Vim-mCh-FKBP
FEATURES             Location/Qualifiers
     source          1..6524
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2117..2126
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2123..2830
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2268..2286)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2536..2555
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2873..3196
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     polyA_signal    3312..3433
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3349..3368)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3403..3422
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3440..3895)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3577..3598)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3789..3808
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3922..4026
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4028..4385
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4028..4219
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4051..4071)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4236..4371
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4420..5214
                     /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             4420..5214
                     /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(4474..4493)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5084..5103
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5402..5421)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5446..5493
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5822..6410
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6311..6330
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atcgccacca tgtccaccag gtccgtgtcc tcgtcctcct accgcaggat
      661 gttcggcggc ccgggcaccg cgagccggcc gagctccagc cggagctacg tgactacgtc
      721 cacccgcacc tacagcctgg gcagcgcgct gcgccccagc accagccgca gcctctacgc
      781 ctcgtccccg ggcggcgtgt atgccacgcg ctcctctgcc gtgcgcctgc ggagcagcgt
      841 gcccggggtg cggctcctgc aggactcggt ggacttctcg ctggccgacg ccatcaacac
      901 cgagttcaag aacacccgca ccaacgagaa ggtggagcta caggagctga atgaccgctt
      961 cgccaactac atcgacaagg tgcgcttcct ggagcagcag aataagatcc tgctggccga
     1021 gctcgagcag ctcaagggcc aaggcaagtc gcgcctgggg gacctctacg aggaggagat
     1081 gcgggagctg cgccggcagg tggaccagct aaccaacgac aaagcccgcg tcgaggtgga
     1141 gcgcgacaac ctggccgagg acatcatgcg cctccgggag aaattgcagg aggagatgct
     1201 tcagagagag gaagccgaaa acaccctgca atctttcaga caggatgttg acaatgcgtc
     1261 tctggcacgt cttgaccttg aacgcaaagt ggaatctttg caagaagaga ttgccttttt
     1321 gaagaaactc cacgaagagg aaatccagga gctgcaggct cagattcagg aacagcatgt
     1381 ccaaatcgat gtggatgttt ccaagcctga cctcacggct gccctgcgtg acgtacgtca
     1441 gcaatatgaa agtgtggctg ccaagaacct gcaggaggca gaagaatggt acaaatccaa
     1501 gtttgctgac ctctctgagg ctgccaaccg gaacaatgac gccctgcgcc aggcaaagca
     1561 ggagtccact gagtaccgga gacaggtgca gtccctcacc tgtgaagtgg atgcccttaa
     1621 aggaaccaat gagtccctgg aacgccagat gcgtgaaatg gaagagaact ttgccgttga
     1681 agctgctaac taccaagaca ctattggccg cctgcaggat gagattcaga atatgaagga
     1741 ggaaatggct cgtcaccttc gtgaatacca agacctgctc aatgttaaga tggcccttga
     1801 cattgagatt gccacctaca ggaagctgct ggaaggcgag gagagcagga tttctctgcc
     1861 tcttccaaac ttttcctccc tgaacctgag ggaaactaat ctggattcac tccctctggt
     1921 tgatacccac tcaaaaagga cacttctgat taagacggtt gaaactagag atggacaggt
     1981 tatcaacgaa acttctcagc atcacgatga ccttgaaccg ggaggcggtg gctcaggcgg
     2041 aggagggtca ggagggggag gcagtggcgg gggcggctct ggaggagggg gttctggtgg
     2101 tggtagcgca ccggtcgcca ccatggtgag caagggcgag gaggataaca tggccatcat
     2161 caaggagttc atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga
     2221 gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt
     2281 gaccaagggt ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg
     2341 ctccaaggcc tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc
     2401 cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac
     2461 ccaggactcc tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa
     2521 cttcccctcc gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga
     2581 gcggatgtac cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa
     2641 ggacggcggc cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca
     2701 gctgcccggc gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta
     2761 caccatcgtg gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga
     2821 gctgtacaag ggcggtggct ctggcggtgg cagtggggga gggggaagtg gaggagtgca
     2881 ggtggaaacc atctccccag gagacgggcg caccttcccc aagcgcggcc agacctgcgt
     2941 ggtgcactac accgggatgc ttgaagatgg aaagaaattt gattcctccc gggacagaaa
     3001 caagcccttt aagtttatgc taggcaagca ggaggtgatc cgaggctggg aagaaggggt
     3061 tgcccagatg agtgtgggtc agagagccaa actgactata tctccagatt atgcctatgg
     3121 tgccactggg cacccaggca tcatcccacc acatgccact ctcgtcttcg atgtggagct
     3181 tctaaaactg gaataagcga ctctagatca taatcagcca taccacattt gtagaggttt
     3241 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
     3301 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     3361 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     3421 tcaatgtatc ttaaggcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     3481 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     3541 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     3601 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     3661 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     3721 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     3781 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     3841 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt
     3901 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     3961 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     4021 aagagtcctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt
     4081 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     4141 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4201 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     4261 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     4321 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     4381 gcaaagatcg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     4441 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     4501 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     4561 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta
     4621 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     4681 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     4741 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     4801 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     4861 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     4921 gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc
     4981 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     5041 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     5101 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     5161 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     5221 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     5281 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     5341 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa
     5401 ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac
     5461 agaataaaac gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct
     5521 ggcactctgt cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct
     5581 tttccccacc ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg
     5641 cggcaggccc tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc
     5701 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     5761 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     5821 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     5881 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     5941 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     6001 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     6061 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     6121 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     6181 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     6241 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     6301 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     6361 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     6421 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     6481 cgttatcccc tgattctgtg gataaccgta ttaccgccat gcat
//
