LOCUS       sequence-385231-        5630 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of mEGFP-HMGB1 WT IDR Fusion Protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5630)
  AUTHORS   Mensah MA, Niskanen H, Magalhaes AP, Basu S, Kircher M, Sczakiel HL,
            Reiter AMV, Elsner J, Meinecke P, Biskup S, Chung BHY, Dombrowsky G,
            Eckmann-Scholz C, Hitz MP, Hoischen A, Holterhus PM, Hulsemann W, 
            Kahrizi K, Kalscheuer VM, Kan A, Krumbiegel M, Kurth I, Leubner J, 
            Longardt AC, Moritz JD, Najmabadi H, Skipalova K, Snijders Blok L, 
            Tzschach A, Wiedersberg E, Zenker M, Garcia-Cabau C, Buschow R, 
            Salvatella X, Kraushar ML, Mundlos S, Caliebe A, Spielmann M, Horn 
            D, Hnisz D
  TITLE     Aberrant phase separation and nucleolar dysfunction in rare genetic 
            diseases.
  JOURNAL   Nature. 2023 Feb;614(7948):564-571. doi: 10.1038/s41586-022-05682-1.
            Epub 2023 Feb 8.
  PUBMED    36755093
REFERENCE   2  (bases 1 to 5630)
  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/s41586-022-05682-1"; journalName: "Nature"; date: 
            "2023-02"; volume: "614"; issue: "7948"; pages: "564-571"
COMMENT     Sequence Label: pRK5-mEGFP-HMGB1-WT-IDR
FEATURES             Location/Qualifiers
     source          1..5630
                     /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      933..942
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(939..941,942..944,945..1652)
                     /codon_start=1
                     /product="enhanced GFP (EGFP) derivative that exhibits 
                     robust folding and enhanced photostability (Zhong et al., 
                     2019)"
                     /label=SiriusGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELY"
     primer_bind     complement(984..1005)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     1592..1613
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      1930..1939
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1943..2077
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1980..1999)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2034..2053
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2146..2503
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2146..2337
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2169..2189)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2354..2489
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2523..2540)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2523..2539)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2532..2554)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2752..3207
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2839..2858)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3049..3070
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3224..3243)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3343..3365
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3403..3421)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3489..3593
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3594..3662,3663..4454)
                     /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(3812..3831)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4625..5213
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5114..5133
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5367..5384
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5501..5522
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5537..5542,5543..5560,5561..5567)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5575..5591
                     /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     5580..5602
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5599..5615
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5599..5615
                     /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 gtgccaccat ggtgagcaag ggcgaggagc
      961 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     1021 tcagcgtgcg cggcgagggc gagggcgatg ccaccaacgg caagctgacc ctgaagttca
     1081 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     1141 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     1201 ccatgcccga aggctacgtc caggagcgca ccatctcctt caaggacgac ggcacctaca
     1261 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     1321 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aacttcaaca
     1381 gccacaacgt ctatatcacg gccgacaagc agaagaacgg catcaaggcg aacttcaaga
     1441 tccgccacaa cgtcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     1501 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccaagc
     1561 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     1621 ccgggatcac tctcggcatg gacgagctgt acacggcgaa cactgctgca gatgacaagc
     1681 agccttatga aaagaaggct gcgaagctga aggaaaaata tgaaaaggat attgctgcat
     1741 atcgagctaa aggaaagcct gatgcagcaa aaaagggagt tgtcaaggct gaaaaaagca
     1801 agaaaaagaa ggaagaggag gaagatgagg aagatgaaga ggatgaggag gaggaggaag
     1861 atgaagaaga tgaagatgaa gaagaagatg atgatgatga ataatctaga gtcgacctgc
     1921 agaagcttgg ccgccatggc ccaacttgtt tattgcagct tataatggtt acaaataaag
     1981 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     2041 gtccaaactc atcaatgtat cttatcatgt ctggatcggg aattaattcg gcgcagcacc
     2101 atggcctgaa ataacctctg aaagaggaac ttggttaggt accttctgag gcggaaagaa
     2161 ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     2221 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     2281 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     2341 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     2401 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     2461 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctgt taacagcttg
     2521 gcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat
     2581 cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat
     2641 cgcccttccc aacagttgcg cagcctgaat ggcgaatggc gcctgatgcg gtattttctc
     2701 cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct
     2761 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     2821 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     2881 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     2941 ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg ccatcgccct
     3001 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     3061 tccaaactgg aacaacactc aaccctatct cgggctattc ttttgattta taagggattt
     3121 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     3181 ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc tgctctgatg
     3241 ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     3301 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     3361 agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat
     3421 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
     3481 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     3541 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     3601 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     3661 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     3721 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     3781 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     3841 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     3901 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     3961 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     4021 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     4081 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     4141 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     4201 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     4261 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     4321 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     4381 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     4441 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     4501 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     4561 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     4621 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     4681 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     4741 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     4801 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     4861 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     4921 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     4981 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     5041 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     5101 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     5161 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     5221 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     5281 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     5341 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     5401 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     5461 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     5521 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     5581 gcggataaca atttcacaca ggaaacagct atgacatgat tacgaattaa
//
