LOCUS       sequence-407457-        8880 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mEGFP-tagged fusion protein, PIEZO1_CBFA2T3 from 
            patient-derived sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8880)
  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 8880)
  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_PIEZO1_CBFA2T3
FEATURES             Location/Qualifiers
     source          1..8880
                     /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(3641..4351,4352..4354,4355..4357))
                     /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(3683..3704)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     4032..4051
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4291..4312
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      4354..4363
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(4439..4642)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4468..4488)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4643..4946
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     CDS             complement(5203..5247)
                     /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    5432..5665
                     /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    6025..6142
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_feature    6634..6759
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             6806..7063
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     enhancer        7065..7444
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7475..7579
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7580..7648,7649..8440)
                     /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(7798..7817)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(8611..8880,1..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 ccggttcagc
     1801 gtggtactgt atctaacgga cctggagggg aaggcgagcc gggacggcta ggaccggctg
     1861 agcccgcttc tgaaggagag gcagctccta ccggtaaaga gggacccaga gaatgagccg
     1921 cttctggcgg acctggaacg ggatcggcaa ctaccgccgt aggaccctga agagattggc
     1981 cgcatacatg atgatgtttc tcccaatcac gatgctgaca gaaggaaccg caatagcgcg
     2041 cagcattaca tcccgagcac gtttcgcttg ctttgcgacc acaattccag catgattcag
     2101 aactgtcttc ttgctggtta attactgtta aagcatcctc gctagcctgg cgtttagcct
     2161 cagccagcgc gcgctccatt ttagcacgct ctgtcgtaat aagttcgtga gccttacgtt
     2221 cagcatcact cacagccttc tgaagttctg acatagcttg acgcttgacc tcattgacag
     2281 cttcttcagc tttgcgccaa atatcctccg ggacataccc agtcagcgta cgtggcagga
     2341 attcacgcgg gacatccagc tgcggacctt cgggtccagc agaagaggaa cgaggacgcg
     2401 ctgctgcagg tgccgggccc ttttttgtat cctctgcatc actgtaacgg cgtgcccaat
     2461 gattcaactc ttcacggtcc gcctcttgac agcgacgcag gacggtcaac gagcggcgtg
     2521 ttttttctac catatccata atacaattta acaagttatt aaggtgcttc cactcctccg
     2581 cccattcgcg ctccgtcagt ttgtgatcaa tcacctcctc ttgacgacta ccaggcacga
     2641 caagcgggcg gtggcgctca cgtaattcac ggggatcggg atgacgatat gcatcacgaa
     2701 aatgatgcgc catcgcaatg tcctccaggc gataatgcgg tggaggagta ggttgtgggg
     2761 gaccattgga tgggctataa cgttgggccg gattcaatgt gcaagggcgc tttgataaat
     2821 gctccgggtg taacgggtca cggtcggagc cgttttcctt cgtacggtca ggagtgcggc
     2881 gttttccatt ttcgttaact tcaagcaaca gctcactgga atcaatcggc gacgaggccg
     2941 atgcatccaa aagcagctgt tcatgctgag ctaaatactg ggctggggtc tgtttggcta
     3001 agcgggcgca atgaagtaat tcacgttgca ataatggaag gttagcctta aggaaaggga
     3061 ttacgaacgg acgcaatggg aagttggtag cctcttgaag tttactgtga aattcctcga
     3121 ttgtaagagt gctatttaca aggcccaaga ccagagtacg aacacgctcc ccaatctctg
     3181 ggctaatgtc cgacccaaat tgttgtaaag tcgtaagaaa gcgtttcaac ttggataact
     3241 ggcgcgcacc acacgcaggg ggtaaatgtt gggtagacag ggaagcggta ctcgaggtcg
     3301 ccggcccgtt cgaaaagccg ttgggtgtac agggcgcgcc attgatagcc gtcggactgt
     3361 gcgaagatcc gttcataagg gtgtgcggtg taaatgaggg agggcgagta gctccttgag
     3421 aggccgccgg gggagggggt ggcatactgg gcggggtact gcgaggctga gtttttactt
     3481 cagctgggct gtctggcata gcactcgctt tagctttgcg atctacggga gccgctaaca
     3541 acgcgcacgg cagaagcagc cagtacagaa ctgcccccag cacgtgaggc tccattcccc
     3601 gggaagcggc cgcacttccc ccggagcctc cgcttccgcc cttgtacagc tcgtccatgc
     3661 cgagagtgat cccggcggcg gtcacgaact ccagcaggac catgtgatcg cgcttctcgt
     3721 tggggtcttt gctcagcttg gactgggtgc tcaggtagtg gttgtcgggc agcagcacgg
     3781 ggccgtcgcc gatgggggtg ttctgctggt agtggtcggc gagctgcacg ctgccgtcct
     3841 cgatgttgtg gcggatcttg aagttcacct tgatgccgtt cttctgcttg tcggccatga
     3901 tatagacgtt gtggctgttg tagttgtact ccagcttgtg ccccaggatg ttgccgtcct
     3961 ccttgaagtc gatgcccttc agctcgatgc ggttcaccag ggtgtcgccc tcgaacttca
     4021 cctcggcgcg ggtcttgtag ttgccgtcgt ccttgaagaa gatggtgcgc tcctggacgt
     4081 agccttcggg catggcggac ttgaagaagt cgtgctgctt catgtggtcg gggtagcggc
     4141 tgaagcactg cacgccgtag gtcagggtgg tcacgagggt gggccagggc acgggcagct
     4201 tgccggtggt gcagatgaac ttcagggtca gcttgccgta ggtggcatcg ccctcgccct
     4261 cgccggacac gctgaacttg tggccgttta cgtcgccgtc cagctcgacc aggatgggca
     4321 ccaccccggt gaacagctcc tcgcccttgc tcaccatggt ggcgaccggt ggatcccggg
     4381 cccgcggtac cgtcgactgc agaattcggt tcactaaacg agcccggttc actaaacgag
     4441 ctctgcttat atagacctcc caccgtacac gcctaccgcc catttgcgtc aatggggcgg
     4501 agttgttacg acattttgga aagtcccgtt gattttggtg ccaaaacaaa ctcccattga
     4561 cgtcaatggg gtggagactt ggaaatcccc gtgagtcaaa ccgctatcca cgcccattga
     4621 tgtactgcca aaaccgcatc accatggtaa tagcgatgac taatacgtag atgtactgcc
     4681 aagtaggaaa gtcccataag gtcatgtact gggcataatg ccaggcgggc catttaccgt
     4741 cattgacgtc aatagggggc gtacttggca tatgatacac ttgatgtact gccaagtggg
     4801 cagtttaccg taaatactcc acccattgac gtcaatggaa agtccctatt ggcgttacta
     4861 tgggaacata cgtcattatt gacgtcaatg ggcgggggtc gttgggcggt cagccaggcg
     4921 ggccatttac cgtaagttat gtaacgcgga actccatata tgggctatga actaatgacc
     4981 ccgtaattga ttactattaa taactagcac gcgttcgaag cttgagctcg gtcccctcgg
     5041 ggttgggagg tgggtctgaa acgataatgg tgaatatccc tgcctaactc tattcactat
     5101 agaaagtaca gcaaaaacta ttcttaaacc taccaagcct cctactatca ttatgaataa
     5161 ttttatatac cacagccaat ttgttatgtt aaaccaattc cacaaacttg cccatttatc
     5221 taattccaat aattcttgtt cattcttttc ttgctggttt tgcgattctt caattaagga
     5281 gtgtattaag cttgtgtaat tgttaatttc tctgtcccac tccatccagg tcgtgtgatt
     5341 ccaaatctgt tccagagatt tattactcca actagcattc caaggcacag cagtggtgca
     5401 aatgagtttt ccagagcaac cccaaatccc caggagctgt tgatccttta ggtatctttc
     5461 cacagccagg attcttgcct ggagctgctt gatgccccag actgtgagtt gcaacagatg
     5521 ctgttgcgcc tcaatagccc tcagcaaatt gttctgctgc tgcactatac cagacaataa
     5581 ttgtctggcc tgtaccgtca gcgtcattga cgctgcgccc atagtgcttc ctgctgctcc
     5641 caagaaccca aggaacaaag ctcctattcc cactgctctt ttttctctct gcaccactct
     5701 tctctttgcc ttggtgggtg ctactcctaa tggttcaatt tttactactt tatatttata
     5761 taattcactt ctccaattgt ccctcatatc tcctccttat ggtgttttac taaacttttc
     5821 catgttctaa tcctcatcct gtctacttgc cacacaatca tcacctgcca tctgttttcc
     5881 ataatcccta atgatctttg cttttcttct tggcactact tttatgtcac tattatcttg
     5941 tattactact gccccttcac ctttccagag gagctttgct ggtcctttcc aaagtggatt
     6001 tctgctgtcc ctgtaataaa cccgaaaatt ttgaattttt gtaatttgtt tttgtaattc
     6061 tttagtttgt atgtctgttg ctattatgtc tactattctt tcccctgcac tgtacccccc
     6121 aatcccccct tttcttttaa aattgtggat gaatactgcc atttgtactg ctgtcttaag
     6181 atgttcagcc tgatctctta cctgtcctat aattttcttt aattctttat tcatagattc
     6241 tactactcct tgactttggg gattgtaggg aatttccagg tctgaagatc ctgctgcttg
     6301 ctgtgctttt ttcttacttt tgttttgctc ttcctctatc ttgtctaaag cttccttggt
     6361 gtcttttatc tctatccttt gatgcacaca atagagggtt gctactgtat tatataatga
     6421 tctaagttct tctgatcctg tctgaaggga tggttgtagc tgtcccagta tttgtctaca
     6481 gccttctgat gtttctaaca ggccagtatt aactgcgaat cgttctagct ccctgcttgc
     6541 ccatactata tgttttaatt tatatttttt ctttccccct ggccttaacc gaattttttc
     6601 ccatcgcgat ctaattctcc cccgcttaat actgacgctc tcgcacccat ctctctcctt
     6661 ctagcctccg ctagtcaaaa tttttggcgt actcaccagt cgccgcccct cgcctcttgc
     6721 cgtgcgcgct tcagcaagcc gagtcctgcg tcgagagagc tcctctggtt tccctttcgc
     6781 tttcaagtcc ctgttcgggc gccactgcta gagattttcc acactgacta aaagggtctg
     6841 agggatctct agttaccaga gtcacacaac agacgggcac acactacttg aagcactcaa
     6901 ggcaagcttt attgaggctt aagcagtggg ttccctagtt agccagagag ctcccaggct
     6961 cagatctggt ctaaccagag agacccagta caggcaaaaa gcagctgctt atatgcagga
     7021 tctgagggct cgccactccc cagtcccgcc caggccacgc ctcccatggt aatagcgatg
     7081 actaatacgt agatgtactg ccaagtagga aagtcccata aggtcatgta ctgggcataa
     7141 tgccaggcgg gccatttacc gtcattgacg tcaatagggg gcgtacttgg catatgatac
     7201 acttgatgta ctgccaagtg ggcagtttac cgtaaatact ccacccattg acgtcaatgg
     7261 aaagtcccta ttggcgttac tatgggaaca tacgtcatta ttgacgtcaa tgggcggggg
     7321 tcgttgggcg gtcagccagg cgggccattt accgtaagtt atgtaacgcg gaactccata
     7381 tatgggctat gaactaatga ccccgtaatt gattactatt aataactagt caataatcaa
     7441 tgtcgaccca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     7501 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     7561 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     7621 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     7681 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     7741 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     7801 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     7861 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     7921 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     7981 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     8041 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     8101 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     8161 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     8221 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     8281 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     8341 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     8401 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     8461 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     8521 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     8581 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     8641 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     8701 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     8761 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     8821 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
//
