LOCUS       sequence-400591-        4700 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Nucleocytoplasmic transport probe.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4700)
  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 4700)
  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 27kDa
FEATURES             Location/Qualifiers
     source          1..4700
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        30..333
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        334..537
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     488..508
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      576..585
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             588..608
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(624..626,627..1340)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(624..626,627..1337)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(666..687)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(927..946)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1274..1295
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1352..1378
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1391..1408
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1461..1479)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1475..1522
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     1548..1567
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1585..2040)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1722..1743)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1934..1953
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2067..2171
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2173..2530
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2173..2364
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2196..2216)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2381..2516
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2565..3359
                     /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             2565..3359
                     /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(2619..2638)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3229..3248
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3547..3566)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3591..3638
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3967..4555
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4456..4475
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
       61 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      121 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      181 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      241 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      301 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      361 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      421 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      481 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctggt
      541 ttagtgaacc gtcagatccg ctagcgctac cggtcgccac catgggccca aaaaagaaaa
      601 gaaaagttgg aggcgggggc agcgtgagca agggcgagga gctgttcacc ggggtggtgc
      661 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
      721 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
      781 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
      841 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
      901 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
      961 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     1021 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     1081 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     1141 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     1201 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     1261 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     1321 tggacgagct gtacaagtaa gtcgacccgg gtatccgtat gatgtgcctg actacgcatg
     1381 atatctcgag caccaccacc accaccactg agatccggct gctaacaaag cccgaaagga
     1441 agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa
     1501 acgggtcttg aggggttttt tgctgaaagg aggaactata tccggatgtg gtttgtccaa
     1561 actcatcaat gtatcttaac gcgtaaattg taagcgttaa tattttgtta aaattcgcgt
     1621 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     1681 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     1741 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     1801 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     1861 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     1921 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     1981 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     2041 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     2101 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     2161 aaaggaagag tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg
     2221 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     2281 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     2341 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     2401 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     2461 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     2521 cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     2581 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     2641 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     2701 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaagac gaggcagcgc
     2761 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     2821 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     2881 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     2941 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     3001 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     3061 cgccagccga actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg
     3121 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     3181 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     3241 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     3301 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     3361 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     3421 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     3481 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag
     3541 gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa
     3601 aagacagaat aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca
     3661 gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc
     3721 ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt
     3781 cggggcggca ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa
     3841 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     3901 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     3961 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     4021 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     4081 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     4141 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     4201 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     4261 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     4321 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     4381 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     4441 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     4501 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     4561 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     4621 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gccatgcatt agttattaat
     4681 agtaatcaat tacggggtca
//
