LOCUS       sequence-471739-        8759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For overexpression of mEGFP-NUP98-DDX10.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8759)
  AUTHORS   Zhang Y, Stoppelkamp I, Fernandez-Pernas P, Allram M, Charman M, 
            Magalhaes AP, Piedavent-Salomon M, Sommer G, Sung YC, Meyer K, Grams
            N, Halko E, Dongre S, Meierhofer D, Malszycki M, Ilik IA, Aktas T, 
            Kraushar ML, Vastenhouw N, Weitzman MD, Grebien F, Niskanen H, Hnisz
            D
  TITLE     Probing condensate microenvironments with a micropeptide killswitch.
  JOURNAL   Nature. 2025 Jun 4. doi: 10.1038/s41586-025-09141-5.
  PUBMED    40468084
REFERENCE   2  (bases 1 to 8759)
  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: "Nature. 
            2025 Jun 4. doi: 10.1038/s41586-025-09141-5."
COMMENT     Sequence Label: pRK5_mEGFP-NUP98-DDX10
FEATURES             Location/Qualifiers
     source          1..8759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     828..845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      934..943
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(940..942,943..945,946..1656)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(985..1006)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1246..1265)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1593..1614
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             complement(4754..4780)
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     regulatory      5059..5068
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    5072..5206
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5109..5128)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5163..5182
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        5275..5632
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5275..5466
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5298..5318)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5483..5618
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5652..5669)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5652..5668)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5661..5683)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5881..6336
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5968..5987)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6178..6199
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6353..6372)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6472..6494
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6532..6550)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6618..6722
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6723..6791,6792..7583)
                     /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     complement(6941..6960)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7754..8342
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8243..8262
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8496..8513
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8630..8651
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8666..8671,8672..8689,8690..8696)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8704..8720
                     /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)."
     primer_bind     8709..8731
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8728..8744
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8728..8744
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgattga attcgtaccg gtcgccacca tggtgagcaa gggcgaggag
      961 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1021 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     1081 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     1141 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1201 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     1261 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1321 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     1381 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     1441 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     1501 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag
     1561 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     1621 gccgggatca ctctcggcat ggacgagctg tacaagaccg gcggcatgtt taacaaatca
     1681 tttggaacac cctttggggg tggcacaggt ggctttggca caacttcaac atttggacag
     1741 aatactggct ttggcactac tagtggaggg gcatttggaa catctgcatt tggttctagc
     1801 aacaatactg gaggcctctt tggaaattca cagactaaac caggaggatt gtttggaacc
     1861 agttcattta gccagccagc tacctccaca agcactggct ttgggtttgg tacgtcaaca
     1921 ggaacagcaa ataccttgtt tggaactgca agcacaggga ccagtctctt ctcatcccaa
     1981 aacaatgcct ttgcacaaaa taaaccaact ggctttggca attttggaac cagtactagc
     2041 agtggaggac tctttggaac cacaaatacc acctctaatc cttttggcag cacatctggc
     2101 tccctctttg ggccaagtag ttttacagct gctcctactg ggactactat taaatttaac
     2161 cctccaactg gtacagatac tatggtcaaa gctggagtta gcactaacat aagtaccaag
     2221 caccagtgta ttactgctat gaaagaatat gaaagcaagt cactagagga acttcgttta
     2281 gaggattatc aggctaacag gaagggccca cagaaccagg tgggagcagg taccacaact
     2341 ggcttgtttg ggtcttctcc agccacttcc agcgcaacag gactcttcag ctcctccacc
     2401 actaattcag gctttgcata tggtcagaac aaaactgcct ttggaactag tacaactgga
     2461 tttggaacaa atccaggtgg tctctttggc caacagaatc agcagactac cagcctcttc
     2521 agcaaaccat ttggccaggc tacaaccacc cagaacactg gcttttcctt tggtaatacc
     2581 agcaccatag gacagccaag caccaacacc atgggattat ttggagtaac ccaagcctca
     2641 cagcctggag gtctttttgg gacagctaca aacaccagca ctgggacagc atttggaaca
     2701 ggaacaggtc tctttgggca gaccaatact ggatttggtg ctgttggttc gaccctgttt
     2761 ggcaataaca agcttactac atttggaagc agcacaacca gtgcaccttc atttggtaca
     2821 accagtggcg ggctctttgg ttttggcaca aataccagtg ggaatagtat ttttggaagt
     2881 aaaccagcac ctgggactct tggaactggg cttggtgcag gatttggaac agctcttggt
     2941 gctggacagg catctttgtt tgggaacaac caacctaaga ttggagggcc tcttggtaca
     3001 ggagcctttg gggcccctgg atttaatact acgacagcca ctttgggctt tggagccccc
     3061 caggccccag tagttcttga tgaagcagat agaatcttgg atatgggctt tgctgatacc
     3121 atgaatgctg ttattgaaaa tctccccaag aaacgtcaga ctttactttt ctcagcaaca
     3181 caaactaaat ctgtaaagga ccttgcacgc ttgagtttga aaaaccctga gtatgtctgg
     3241 gttcatgaaa aagcaaaata tagcacccct gccactttgg aacagaacta catagtctgt
     3301 gagctgcagc aaaaaataag tgtgctgtat tcctttttga gaagccatct gaagaagaag
     3361 agcattgtat ttttttccag ttgcaaagag gtccagtatc tgtaccgagt gttttgccgg
     3421 ctacgtcctg gtgtttctat ccttgcactc catggtcgac agcagcaaat gagaagaatg
     3481 gaagtctata atgagtttgt ccgtaagaga gctgcagtac tctttgctac tgatattgca
     3541 gccaggggtc tggatttccc ggccgtgaat tgggttcttc agtttgattg tcctgaggat
     3601 gccaacacat atattcacag agcaggtaga actgccaggt acaaagagga tggtgaagct
     3661 ttgctaattt tgctaccctc agaaaaagct atggtgcagc agcttcttca gaagaaagta
     3721 cctgtgaagg aaatcaaaat caatccagaa aaacttatag atgtccagaa aaaattggaa
     3781 tctattttag ctcaagatca agatttaaaa gaaagagctc aaaggtgttt cgtctcctat
     3841 gtacgatctg tatatctgat gaaggataaa gaagtatttg atgtgagcaa gttacctata
     3901 cctgaatatg ccctgtctct tggtcttgct gtggcaccac gcgtaagatt tcttcagaaa
     3961 atgcagaaac aacccaccaa agaattggta aggagccaag ccgataaagt aattgagcca
     4021 agggctccct ccctcaccaa tgacgaagtg gaagaattta gagcctactt caatgagaaa
     4081 atgtccatcc ttcaaaaagg tggaaaaaga ctcgaaggga cagagcacag acaggataat
     4141 gatactggta atgaagaaca ggaagaagaa gaagacgatg aagaagaaat ggaagagaaa
     4201 ctggcaaaag caaaaggatc tcaagcccca tctcttccta acaccagtga ggcacagaag
     4261 atcaaggaag ttcctacaca gttcttggac agagatgagg aggaagaaga tgctgatttc
     4321 ttgaaggtga agcggcataa tgtgtttgga ttggacctta aagacgagaa aacattacag
     4381 aagaaagaac cttctaaatc cagcatcaag aaaaaaatga ccaaagttgc agaagcaaaa
     4441 aaagtaatga agagaaattt taaagtgaat aagaagataa catttactga tgaaggggag
     4501 ttggttcagc agtggccaca aatgcagaaa tctgccatca aggatgctga ggaagatgat
     4561 gacacaggtg gtatcaactt acataaagca aaggaaagac ttcaggaaga ggacaaattt
     4621 gacaaagaag aatataggaa aaaaattaag gcaaagcatc gggagaaaag actgaaagaa
     4681 agggaagcca gaagagaagc caacaagaga caagcaaagg ccaaagatga agaggaagcc
     4741 tttctggatt ggagtgatga tgatgatgat gatgatgatg gatttgatcc aagcacactc
     4801 ccagatccag ataaatacag aagctctgaa gattcagata gtgaagatat ggaaaataaa
     4861 ataagtgata ccaagaagaa gcaggggatg aagaagagga gcaacagtga agtggaagac
     4921 gtgggaccaa caagtcataa cagaaagaag gccaggtggg acactttaga gcctttggat
     4981 accggcctgt ctttagcaga ggatgaagag ctggtgttac atctgctaag aagtcaaagc
     5041 taaacctgca gaagcttggc cgccatggcc caacttgttt attgcagctt ataatggtta
     5101 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     5161 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggatcggga attaattcgg
     5221 cgcagcacca tggcctgaaa taacctctga aagaggaact tggttaggta ccttctgagg
     5281 cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     5341 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     5401 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     5461 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     5521 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
     5581 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctgtt
     5641 aacagcttgg cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     5701 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     5761 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggcg cctgatgcgg
     5821 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt
     5881 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     5941 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     6001 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     6061 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
     6121 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     6181 gactcttgtt ccaaactgga acaacactca accctatctc gggctattct tttgatttat
     6241 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     6301 acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct
     6361 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     6421 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     6481 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     6541 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     6601 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     6661 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     6721 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     6781 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     6841 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     6901 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     6961 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     7021 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     7081 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     7141 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     7201 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     7261 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     7321 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     7381 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     7441 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     7501 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     7561 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     7621 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     7681 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     7741 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     7801 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     7861 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     7921 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     7981 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     8041 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     8101 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     8161 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     8221 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     8281 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     8341 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     8401 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
     8461 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     8521 aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct
     8581 ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt
     8641 agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg
     8701 gaattgtgag cggataacaa tttcacacag gaaacagcta tgacatgatt acgaattaa
//
