LOCUS       sequence-400072-        4628 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Twist mutant for nucleocytoplasmic transport studies.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4628)
  AUTHORS   Andreu I, Granero-Moya I, Chahare NR, Clein K, Molina-Jordan M, 
            Beedle AEM, Elosegui-Artola A, Abenza JF, Rossetti L, Trepat X, 
            Raveh B, Roca-Cusachs P
  TITLE     Mechanical force application to the nucleus regulates 
            nucleocytoplasmic transport.
  JOURNAL   Nat Cell Biol. 2022 Jun;24(6):896-905. doi: 
            10.1038/s41556-022-00927-7. Epub 2022 Jun 9.
  PUBMED    35681009
REFERENCE   2  (bases 1 to 4628)
  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"; doi: 
            "10.1038/s41556-022-00927-7"; journalName: "Nat Cell Biol"; date: 
            "2022-06"; volume: "24"; issue: "6"; pages: "896-905"
COMMENT     Sequence Label: H_NLS-mutNLS V5-Twist
FEATURES             Location/Qualifiers
     source          1..4628
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        24..327
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        328..531
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     482..502
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      570..579
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             582..602
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             603..644
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             1274..1300
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1313..1330
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1383..1401)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1397..1444
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     1470..1489
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1507..1962)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1644..1665)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1856..1875
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1989..2093
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2095..2452
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2095..2286
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2118..2138)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2303..2438
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2365..2384
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2487..3281
                     /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             2487..3281
                     /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(2541..2560)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3151..3170
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3469..3488)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3513..3560
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3889..4477
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4378..4397
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
       61 ccgcccaacg acccacgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
      121 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
      181 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
      241 cccgcctggc actatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
      301 tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt
      361 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
      421 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
      481 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg
      541 aaccgtcaga tccgctagcg ctaccggtcg ccaccatggg cccaaaaaag aaaagaaaag
      601 ttggaaagcc tattcctaat cctcttctag gtctagattc tactatgcag gacgtgtcca
      661 gctcgccagt ctctccggcc gacgacagcc tgagcaacag cgaggaggag ccggaccggc
      721 agcagccggc gagcggcaag cgcggggctc gcaggagacg cagcagtcgg cgcagcgcgg
      781 gcggcagcgc ggggcccggc ggggccacgg gcgggggcat cggaggcggc gacgagccag
      841 gcagcccggc ccagggcagg cgcggcaaga aatctgcggg cggaggcggc ggcggcggcg
      901 cgggcggagg tggtggcggc ggcggcggca gcagcagcgg gggcgggagc ccgcagtcgt
      961 acgaggagct gcagacccag cgggtcatgg ctaacgtgcg ggagcgccag cgcacgcagt
     1021 cgctgaacga ggcgttcgcc gccctgcgca agatcatccc cacgctgccc tcggacaagc
     1081 tgagcaagat tcagaccctc aaactggcgg ccaggtacat cgacttcctg taccaggtcc
     1141 tgcagagcga cgagctggac tccaagatgg caagctgcag ctatgtggcc cacgagcggc
     1201 tcagctacgc cttctccgtc tggaggatgg agggggcctg gtccatgtcc gcgtcccact
     1261 aggtcgaccc gggtatccgt atgatgtgcc tgactacgca tgatatctcg agcaccacca
     1321 ccaccaccac tgagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc
     1381 caccgctgag caataactag cataacccct tggggcctct aaacgggtct tgaggggttt
     1441 tttgctgaaa ggaggaacta tatccggatg tggtttgtcc aaactcatca atgtatctta
     1501 acgcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     1561 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     1621 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     1681 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     1741 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     1801 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgagg aaggaaggga
     1861 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     1921 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg
     1981 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2041 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg
     2101 cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     2161 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     2221 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     2281 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     2341 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcccctga
     2401 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc
     2461 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc
     2521 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     2581 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     2641 acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca
     2701 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     2761 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     2821 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     2881 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     2941 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     3001 ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     3061 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     3121 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
     3181 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
     3241 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
     3301 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     3361 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     3421 cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa
     3481 ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca
     3541 cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga
     3601 taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca
     3661 ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc
     3721 catagcctca ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     3781 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     3841 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     3901 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     3961 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     4021 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     4081 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     4141 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     4201 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     4261 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     4321 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     4381 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     4441 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     4501 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     4561 ttctgtggat aaccgtatta ccgccatgca ttagttatta atagtaatca attacggggt
     4621 cattagtt
//
