LOCUS       sequence-418466-        8874 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mEGFP-tagged fusion protein, EML4_BRAF from patient-derived 
            sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8874)
  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 8874)
  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_EML4_BRAF
FEATURES             Location/Qualifiers
     source          1..8874
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(441..1232,1233..1301))
                     /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     1064..1083
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1302..1406)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        1437..1816
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     LTR             1818..2075
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    2122..2247
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    2739..2856
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_feature    3216..3449
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             3634..3678
                     /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"
     enhancer        3935..4238
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4239..4442
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4393..4413
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      4518..4527
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4524..4526,4527..4529,4530..5240)
                     /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(4569..4590)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4830..4849)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5177..5198
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(7328..7347)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    7393..7448
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(7447..7466)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(7809..7825)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7809..7825)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7822..7844)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7833..7849
                     /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)."
     promoter        complement(join(7857..7863,7864..7881,7882..7887))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7902..7923
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        7982..8177
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      8028..8163
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    8183..8317
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8220..8239)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8274..8293
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(8385..8402)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(8556..8874,1..270))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8636..8655)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
       61 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
      121 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
      181 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
      241 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
      301 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
      361 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
      421 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
      481 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
      541 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
      601 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
      661 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
      721 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
      781 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
      841 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
      901 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
      961 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     1021 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     1081 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     1141 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     1201 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     1261 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     1321 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     1381 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgggtcgaca
     1441 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
     1501 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     1561 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
     1621 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     1681 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     1741 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     1801 catcgctatt accatgggag gcgtggcctg ggcgggactg gggagtggcg agccctcaga
     1861 tcctgcatat aagcagctgc tttttgcctg tactgggtct ctctggttag accagatctg
     1921 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
     1981 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
     2041 cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa
     2101 gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg
     2161 gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag
     2221 aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg
     2281 gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg
     2341 gcaagcaggg agctagaacg attcgcagtt aatactggcc tgttagaaac atcagaaggc
     2401 tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga
     2461 tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac
     2521 accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagaa aaaagcacag
     2581 caagcagcag gatcttcaga cctggaaatt ccctacaatc cccaaagtca aggagtagta
     2641 gaatctatga ataaagaatt aaagaaaatt ataggacagg taagagatca ggctgaacat
     2701 cttaagacag cagtacaaat ggcagtattc atccacaatt ttaaaagaaa aggggggatt
     2761 ggggggtaca gtgcagggga aagaatagta gacataatag caacagacat acaaactaaa
     2821 gaattacaaa aacaaattac aaaaattcaa aattttcggg tttattacag ggacagcaga
     2881 aatccacttt ggaaaggacc agcaaagctc ctctggaaag gtgaaggggc agtagtaata
     2941 caagataata gtgacataaa agtagtgcca agaagaaaag caaagatcat tagggattat
     3001 ggaaaacaga tggcaggtga tgattgtgtg gcaagtagac aggatgagga ttagaacatg
     3061 gaaaagttta gtaaaacacc ataaggagga gatatgaggg acaattggag aagtgaatta
     3121 tataaatata aagtagtaaa aattgaacca ttaggagtag cacccaccaa ggcaaagaga
     3181 agagtggtgc agagagaaaa aagagcagtg ggaataggag ctttgttcct tgggttcttg
     3241 ggagcagcag gaagcactat gggcgcagcg tcaatgacgc tgacggtaca ggccagacaa
     3301 ttattgtctg gtatagtgca gcagcagaac aatttgctga gggctattga ggcgcaacag
     3361 catctgttgc aactcacagt ctggggcatc aagcagctcc aggcaagaat cctggctgtg
     3421 gaaagatacc taaaggatca acagctcctg gggatttggg gttgctctgg aaaactcatt
     3481 tgcaccactg ctgtgccttg gaatgctagt tggagtaata aatctctgga acagatttgg
     3541 aatcacacga cctggatgga gtgggacaga gaaattaaca attacacaag cttaatacac
     3601 tccttaattg aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt attggaatta
     3661 gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg gtatataaaa
     3721 ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc tgtactttct
     3781 atagtgaata gagttaggca gggatattca ccattatcgt ttcagaccca cctcccaacc
     3841 ccgaggggac cgagctcaag cttcgaacgc gtgctagtta ttaatagtaa tcaattacgg
     3901 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
     3961 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
     4021 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
     4081 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
     4141 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
     4201 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
     4261 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
     4321 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
     4381 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
     4441 ctcgtttagt gaaccgggct cgtttagtga accgaattct gcagtcgacg gtaccgcggg
     4501 cccgggatcc accggtcgcc accatggtga gcaagggcga ggagctgttc accggggtgg
     4561 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     4621 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
     4681 agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca
     4741 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     4801 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     4861 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     4921 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     4981 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg
     5041 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     5101 ccgtgctgct gcccgacaac cactacctga gcacccagtc caagctgagc aaagacccca
     5161 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     5221 gcatggacga gctgtacaag ggcggaagcg gaggctccgg gggaagtgcg gccgcttccc
     5281 ggggaatgga tggattcgcc ggtagcttag atgactctat ctccgccgcc tccacttctg
     5341 acgttcagga tcgtctttct gcgctggaaa gtcgcgttca acagcaagag gacgaaatca
     5401 cagtcctgaa agcggcactt gcagatgtct tgcgccgtct ggctatcagt gaggatcacg
     5461 tagcgtcagt caagaaatcg gtatcatcaa agggtcaacc ctctccccgc gcggttattc
     5521 cgatgagctg catcacgaac ggctccggcg ctaatcgtaa gccctctcat acgagtgcgg
     5581 tttccattgc tggcaaagag acattatcct ctgcagcaaa atcaggaacc gagaagaaga
     5641 aggaaaaacc gcaagggcaa cgcgaaaaga aggaggaatc gcactcgaac gatcagtcac
     5701 cccagatccg tgcctcgcct tctccgcagc caagcagtca gccgttacaa atccatcgcc
     5761 agacgccaga aagcaaaaac gccaccccga ctaagagtat taagcgtcct agcccagccg
     5821 aaaaaagcca taactcctgg gaaaatagtg atgattcccg caacaagtta tcgaagattc
     5881 ccagcactcc gaaacttatc ccgaaagtta caaagacggc ggacaaacat aaggatgtga
     5941 tcatcaacca aggatcaaca acgggtctgt cggcgacccc gcctgccagt ctgcctggtt
     6001 cgctgaccaa tgtaaaggca cttcagaagt cacccggccc acaacgcgag cgtaaaagca
     6061 gctcgtctag cgaagaccgt aaccgtatga agaccctggg gcgtcgtgac tcctctgacg
     6121 actgggagat ccccgatggg cagatcaccg tcggacaacg cattggctca gggtcgttcg
     6181 gcactgtgta caagggtaag tggcacgggg atgtagcggt taaaatgtta aatgttactg
     6241 ctccgactcc acaacagtta caggctttca aaaatgaggt aggagtgttg cgcaaaactc
     6301 gtcacgtcaa tatcctttta tttatggggt acagtactaa gccgcagctt gcaatcgtta
     6361 cacaatggtg cgaaggatcg tcactttatc accatttgca catcattgag acaaagtttg
     6421 aaatgatcaa gctgatcgat attgcacgtc agaccgcgca aggaatggat tacttacatg
     6481 caaagtcaat cattcatcgt gaccttaaat caaacaatat ctttcttcac gaggacttga
     6541 ctgttaagat tggcgacttt gggctggcaa ctgtgaagtc tcgttggtct ggctcacatc
     6601 aatttgaaca attaagcggc tcgatccttt ggatggctcc agaggtgatt cgcatgcaag
     6661 acaaaaatcc gtattcattt caatcggatg tgtacgcgtt cggaattgtg ctgtacgagt
     6721 taatgaccgg ccagcttccg tactcgaaca tcaacaatcg cgatcaaatt atttttatgg
     6781 tcggtcgcgg atacttgtct ccggatctgt ccaaagtccg tagtaattgc ccaaaggcaa
     6841 tgaaacgtct tatggcagag tgtttgaaga aaaaacgtga cgaacgtccc ttgtttccac
     6901 agattcttgc gtcaatcgag cttcttgcgc gtagtttacc aaagattcac cgctcagcgt
     6961 ctgaaccttc acttaatcgc gcaggtttcc aaactgaaga cttctccttg tacgcctgtg
     7021 cgagccccaa gacaccaatc caggccggcg gttacggcga atttgcggct tttaaatgaa
     7081 ccggttctag ataagattac ggtaccttta agaccaatga cttacaaggc agctgtagat
     7141 cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc ccaaagaaga
     7201 caagatctgc tttttgcctg tactgggtct ctctggttag accagatctg agcctgggag
     7261 ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttcag ctgctcgagc
     7321 tagcagatct ttttccctct gccaaaaatt atggggacat catgaagccc cttgagcatc
     7381 tgacttctgg ctaataaagg aaatttattt tcattgcaat agtgtgttgg aattttttgt
     7441 gtctctcact cggaaggaca tatgggaggg caaatcattt aaaacatcag aatgagtatt
     7501 tggtttagag tttggcaaca tatgcccata tgctggctgc catgaacaaa ggttggctat
     7561 aaagaggtca tcagtatatg aaacagcccc ctgctgtcca ttccttattc catagaaaag
     7621 ccttgacttg aggttagatt ttttttatat tttgttttgt gttatttttt tctttaacat
     7681 ccctaaaatt ttccttacat gttttactag ccagattttt cctcctctcc tgactactcc
     7741 cagtcatagc tgtccctctt ctcttatgga gatccctcga cctgcagccc aagcttggcg
     7801 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     7861 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     7921 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagcggatc
     7981 cgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     8041 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     8101 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     8161 gcctaggctt ttgcaaaaag ctaacttgtt tattgcagct tataatggtt acaaataaag
     8221 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     8281 gtccaaactc atcaatgtat cttatcatgt ctggatccgc tgcattaatg aatcggccaa
     8341 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     8401 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     8461 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     8521 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     8581 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     8641 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     8701 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     8761 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     8821 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caac
//
