LOCUS       sequence-385201-        5982 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of mEGFP-HMGB3 WT Fusion Protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5982)
  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 5982)
  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-HMGB3-WT
FEATURES             Location/Qualifiers
     source          1..5982
                     /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      2282..2291
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2295..2429
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2332..2351)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2386..2405
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2498..2855
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2498..2689
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2521..2541)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2706..2841
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2875..2892)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2875..2891)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2884..2906)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3104..3559
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3191..3210)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3401..3422
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3576..3595)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3695..3717
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3755..3773)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3841..3945
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3946..4014,4015..4806)
                     /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(4164..4183)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4977..5565
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5466..5485
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5719..5736
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5853..5874
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5889..5894,5895..5912,5913..5919)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5927..5943
                     /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     5932..5954
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5951..5967
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5951..5967
                     /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 acacggcgat ggctaaaggt gaccccaaga
     1681 aaccaaaggg caagatgtcc gcttatgcct tctttgtgca gacatgcaga gaagaacata
     1741 agaagaaaaa cccagaggtc cctgtcaatt ttgcggaatt ttccaagaag tgctctgaga
     1801 ggtggaagac gatgtccggg aaagagaaat ctaaatttga tgaaatggca aaggcagata
     1861 aagtgcgcta tgatcgggaa atgaaggatt atggaccagc taagggaggc aagaagaaga
     1921 aggatcctaa tgctcccaaa aggccaccgt ctggattctt cctgttctgt tcagaattcc
     1981 gccccaagat caaatccaca aaccccggca tctctattgg agacgtggca aaaaagctgg
     2041 gtgagatgtg gaataattta aatgacagtg aaaagcagcc ttacatcact aaggcggcaa
     2101 agctgaagga gaagtatgag aaggatgttg ctgactataa gtcgaaagga aagtttgatg
     2161 gtgcaaaggg tcctgctaaa gttgcccgga aaaaggtgga agaggaagat gaagaagagg
     2221 aggaagagga ggaggaagaa gaggaagaag aggatgaata aagtcgacct gcagaagctt
     2281 ggccgccatg gcccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     2341 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     2401 tcatcaatgt atcttatcat gtctggatcg ggaattaatt cggcgcagca ccatggcctg
     2461 aaataacctc tgaaagagga acttggttag gtaccttctg aggcggaaag aaccagctgt
     2521 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     2581 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     2641 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     2701 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     2761 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     2821 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct gttaacagct tggcactggc
     2881 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc
     2941 agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc
     3001 ccaacagttg cgcagcctga atggcgaatg gcgcctgatg cggtattttc tccttacgca
     3061 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     3121 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     3181 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     3241 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     3301 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     3361 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     3421 ggaacaacac tcaaccctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     3481 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     3541 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     3601 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     3661 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     3721 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     3781 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     3841 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     3901 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     3961 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     4021 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     4081 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     4141 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     4201 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     4261 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     4321 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     4381 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     4441 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     4501 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     4561 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     4621 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     4681 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     4741 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     4801 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     4861 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     4921 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     4981 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     5041 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     5101 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     5161 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     5221 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     5281 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     5341 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     5401 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     5461 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     5521 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     5581 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     5641 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     5701 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     5761 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     5821 gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca
     5881 ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa
     5941 caatttcaca caggaaacag ctatgacatg attacgaatt aa
//
