LOCUS       sequence-407511-        8001 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mEGFP-tagged fusion protein, EWSR1_WT1 from patient-derived 
            sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8001)
  AUTHORS   Tripathi S, Shirnekhi HK, Gorman SD, Chandra B, Baggett DW, Park CG,
            Somjee R, Lang B, Hosseini SMH, Pioso BJ, Li Y, Iacobucci I, Gao Q, 
            Edmonson MN, Rice SV, Zhou X, Bollinger J, Mitrea DM, White MR, 
            McGrail DJ, Jarosz DF, Yi SS, Babu MM, Mullighan CG, Zhang J, Sahni 
            N, Kriwacki RW
  TITLE     Defining the condensate landscape of fusion oncoproteins.
  JOURNAL   Nat Commun. 2023 Sep 28;14(1):6008. doi: 10.1038/s41467-023-41655-2.
  PUBMED    37770423
REFERENCE   2  (bases 1 to 8001)
  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: "Nat 
            Commun."; date: "2023-09-28"; volume: "14(1):6008. doi"; pages: " 
            10.1038/s41467-023-41655-2"
COMMENT     Sequence Label: CL20_mEGFP_EWSR1_WT1
FEATURES             Location/Qualifiers
     source          1..8001
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     220..239
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     473..490
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    558..692
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(582..601)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     636..655
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(698..893)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      712..847
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     protein_bind    952..973
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(988..993,994..1011,1012..1018)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1026..1042
                     /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     1031..1053
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1050..1066
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1050..1066
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1409..1428
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    1427..1482
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     1528..1547
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(join(2762..3472,3473..3475,3476..3478))
                     /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(2804..2825)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     3153..3172
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3412..3433
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3475..3484
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(3560..3763)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3589..3609)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3764..4067
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     CDS             complement(4324..4368)
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    4553..4786
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    5146..5263
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_feature    5755..5880
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             5927..6184
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     enhancer        6186..6565
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6596..6700
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6701..6769,6770..7561)
                     /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(6919..6938)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(7732..8001,1..319)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
       61 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
      121 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
      181 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
      241 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
      301 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
      361 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
      421 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
      481 agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc
      541 gattcattaa tgcagcggat ccagacatga taagatacat tgatgagttt ggacaaacca
      601 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
      661 ttgtaaccat tataagctgc aataaacaag ttagcttttt gcaaaagcct aggcctccaa
      721 aaaagcctcc tcactacttc tggaatagct cagaggccga ggcggcctcg gcctctgcat
      781 aaataaaaaa aattagtcag ccatggggcg gagaatgggc ggaactgggc ggagttaggg
      841 gcgggatggg cggagttagg ggcgggacta tggttgctga ctaattgaga tgcggatccg
      901 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
      961 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
     1021 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa
     1081 gcttgggctg caggtcgagg gatctccata agagaagagg gacagctatg actgggagta
     1141 gtcaggagag gaggaaaaat ctggctagta aaacatgtaa ggaaaatttt agggatgtta
     1201 aagaaaaaaa taacacaaaa caaaatataa aaaaaatcta acctcaagtc aaggcttttc
     1261 tatggaataa ggaatggaca gcagggggct gtttcatata ctgatgacct ctttatagcc
     1321 aacctttgtt catggcagcc agcatatggg catatgttgc caaactctaa accaaatact
     1381 cattctgatg ttttaaatga tttgccctcc catatgtcct tccgagtgag agacacaaaa
     1441 aattccaaca cactattgca atgaaaataa atttccttta ttagccagaa gtcagatgct
     1501 caaggggctt catgatgtcc ccataatttt tggcagaggg aaaaagatct gctagctcga
     1561 gcagctgaag ctttattgag gcttaagcag tgggttccct agttagccag agagctccca
     1621 ggctcagatc tggtctaacc agagagaccc agtacaggca aaaagcagat cttgtcttct
     1681 ttgggagtga attagccctt ccagtccccc cttttctttt aaaaagtggc taagatctac
     1741 agctgccttg taagtcattg gtcttaaagg taccgtaatc ttatctagaa ccggttcaca
     1801 gggcaagctg cagtttcgtc atattgcgtt gatgcatatt atgatgacgg acaagctcgt
     1861 cagaacgggc gaactttttc tggcaactag gccagcgaca ggagaacggc ttttcccccg
     1921 tgtgcgtacg ggtatgagtc ttcagatgat ccgaacgaga gaatttgcgt tggcaggttt
     1981 tacactgaaa gggcttgaca cctgtatgac ggcgttggtg acgtttaagt tgatccgagc
     2041 gactgaagcg acgttcgcag tctttgaagt cacactggta gggcttctcg ctctgctgtc
     2101 cgtaactaga ggactgctgg ctgtattggg agggtgcctg gctatatgac cccgtctggg
     2161 gcgggtatga tgttggcggc tgttggccgt aagatgattg ttggccgtag ctcgactgct
     2221 gcccgtagct gcttggctga ccgtatgtat tctgttgaga gtacgaagac tggtcatagc
     2281 tagtcggctg agtggatgag taacttgtgg gggcggtcgc agctggctgt tgtccgtaag
     2341 cagggtacgc aggctgagtc ccgtaggcgg actgggctgc atacgatgcc tgagtagtcg
     2401 taacagtggc tgtcgttgtg tcgtaggctc cagttccata cccctgtact ggttgcgagt
     2461 atgcctgagg ggcagttggg gttgtgtacc cggtcggagg ttgcccgtat gatgtcgcgt
     2521 aggcagtttg tccataggtc gcagtggtct gtgcctgagt atacgacaca tctgttggct
     2581 gaccatacgt gccataactt tgctgaccat aagcctgggt tgtttgagca tatccctgcg
     2641 tgggctgagc ggtataggca gaataaccct gctgggccgc cgcttgcgag taggtcgaat
     2701 agtcggtgga agccattccc cgggaagcgg ccgcacttcc cccggagcct ccgcttccgc
     2761 ccttgtacag ctcgtccatg ccgagagtga tcccggcggc ggtcacgaac tccagcagga
     2821 ccatgtgatc gcgcttctcg ttggggtctt tgctcagctt ggactgggtg ctcaggtagt
     2881 ggttgtcggg cagcagcacg gggccgtcgc cgatgggggt gttctgctgg tagtggtcgg
     2941 cgagctgcac gctgccgtcc tcgatgttgt ggcggatctt gaagttcacc ttgatgccgt
     3001 tcttctgctt gtcggccatg atatagacgt tgtggctgtt gtagttgtac tccagcttgt
     3061 gccccaggat gttgccgtcc tccttgaagt cgatgccctt cagctcgatg cggttcacca
     3121 gggtgtcgcc ctcgaacttc acctcggcgc gggtcttgta gttgccgtcg tccttgaaga
     3181 agatggtgcg ctcctggacg tagccttcgg gcatggcgga cttgaagaag tcgtgctgct
     3241 tcatgtggtc ggggtagcgg ctgaagcact gcacgccgta ggtcagggtg gtcacgaggg
     3301 tgggccaggg cacgggcagc ttgccggtgg tgcagatgaa cttcagggtc agcttgccgt
     3361 aggtggcatc gccctcgccc tcgccggaca cgctgaactt gtggccgttt acgtcgccgt
     3421 ccagctcgac caggatgggc accaccccgg tgaacagctc ctcgcccttg ctcaccatgg
     3481 tggcgaccgg tggatcccgg gcccgcggta ccgtcgactg cagaattcgg ttcactaaac
     3541 gagcccggtt cactaaacga gctctgctta tatagacctc ccaccgtaca cgcctaccgc
     3601 ccatttgcgt caatggggcg gagttgttac gacattttgg aaagtcccgt tgattttggt
     3661 gccaaaacaa actcccattg acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa
     3721 accgctatcc acgcccattg atgtactgcc aaaaccgcat caccatggta atagcgatga
     3781 ctaatacgta gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat
     3841 gccaggcggg ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca
     3901 cttgatgtac tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga
     3961 aagtccctat tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt
     4021 cgttgggcgg tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat
     4081 atgggctatg aactaatgac cccgtaattg attactatta ataactagca cgcgttcgaa
     4141 gcttgagctc ggtcccctcg gggttgggag gtgggtctga aacgataatg gtgaatatcc
     4201 ctgcctaact ctattcacta tagaaagtac agcaaaaact attcttaaac ctaccaagcc
     4261 tcctactatc attatgaata attttatata ccacagccaa tttgttatgt taaaccaatt
     4321 ccacaaactt gcccatttat ctaattccaa taattcttgt tcattctttt cttgctggtt
     4381 ttgcgattct tcaattaagg agtgtattaa gcttgtgtaa ttgttaattt ctctgtccca
     4441 ctccatccag gtcgtgtgat tccaaatctg ttccagagat ttattactcc aactagcatt
     4501 ccaaggcaca gcagtggtgc aaatgagttt tccagagcaa ccccaaatcc ccaggagctg
     4561 ttgatccttt aggtatcttt ccacagccag gattcttgcc tggagctgct tgatgcccca
     4621 gactgtgagt tgcaacagat gctgttgcgc ctcaatagcc ctcagcaaat tgttctgctg
     4681 ctgcactata ccagacaata attgtctggc ctgtaccgtc agcgtcattg acgctgcgcc
     4741 catagtgctt cctgctgctc ccaagaaccc aaggaacaaa gctcctattc ccactgctct
     4801 tttttctctc tgcaccactc ttctctttgc cttggtgggt gctactccta atggttcaat
     4861 ttttactact ttatatttat ataattcact tctccaattg tccctcatat ctcctcctta
     4921 tggtgtttta ctaaactttt ccatgttcta atcctcatcc tgtctacttg ccacacaatc
     4981 atcacctgcc atctgttttc cataatccct aatgatcttt gcttttcttc ttggcactac
     5041 ttttatgtca ctattatctt gtattactac tgccccttca cctttccaga ggagctttgc
     5101 tggtcctttc caaagtggat ttctgctgtc cctgtaataa acccgaaaat tttgaatttt
     5161 tgtaatttgt ttttgtaatt ctttagtttg tatgtctgtt gctattatgt ctactattct
     5221 ttcccctgca ctgtaccccc caatcccccc ttttctttta aaattgtgga tgaatactgc
     5281 catttgtact gctgtcttaa gatgttcagc ctgatctctt acctgtccta taattttctt
     5341 taattcttta ttcatagatt ctactactcc ttgactttgg ggattgtagg gaatttccag
     5401 gtctgaagat cctgctgctt gctgtgcttt tttcttactt ttgttttgct cttcctctat
     5461 cttgtctaaa gcttccttgg tgtcttttat ctctatcctt tgatgcacac aatagagggt
     5521 tgctactgta ttatataatg atctaagttc ttctgatcct gtctgaaggg atggttgtag
     5581 ctgtcccagt atttgtctac agccttctga tgtttctaac aggccagtat taactgcgaa
     5641 tcgttctagc tccctgcttg cccatactat atgttttaat ttatattttt tctttccccc
     5701 tggccttaac cgaatttttt cccatcgcga tctaattctc ccccgcttaa tactgacgct
     5761 ctcgcaccca tctctctcct tctagcctcc gctagtcaaa atttttggcg tactcaccag
     5821 tcgccgcccc tcgcctcttg ccgtgcgcgc ttcagcaagc cgagtcctgc gtcgagagag
     5881 ctcctctggt ttccctttcg ctttcaagtc cctgttcggg cgccactgct agagattttc
     5941 cacactgact aaaagggtct gagggatctc tagttaccag agtcacacaa cagacgggca
     6001 cacactactt gaagcactca aggcaagctt tattgaggct taagcagtgg gttccctagt
     6061 tagccagaga gctcccaggc tcagatctgg tctaaccaga gagacccagt acaggcaaaa
     6121 agcagctgct tatatgcagg atctgagggc tcgccactcc ccagtcccgc ccaggccacg
     6181 cctcccatgg taatagcgat gactaatacg tagatgtact gccaagtagg aaagtcccat
     6241 aaggtcatgt actgggcata atgccaggcg ggccatttac cgtcattgac gtcaataggg
     6301 ggcgtacttg gcatatgata cacttgatgt actgccaagt gggcagttta ccgtaaatac
     6361 tccacccatt gacgtcaatg gaaagtccct attggcgtta ctatgggaac atacgtcatt
     6421 attgacgtca atgggcgggg gtcgttgggc ggtcagccag gcgggccatt taccgtaagt
     6481 tatgtaacgc ggaactccat atatgggcta tgaactaatg accccgtaat tgattactat
     6541 taataactag tcaataatca atgtcgaccc aggtggcact tttcggggaa atgtgcgcgg
     6601 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     6661 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     6721 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     6781 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     6841 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     6901 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     6961 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     7021 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     7081 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     7141 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     7201 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     7261 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     7321 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     7381 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     7441 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     7501 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     7561 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     7621 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     7681 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     7741 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     7801 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     7861 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     7921 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     7981 cgataagtcg tgtcttaccg g
//
