LOCUS       sequence-407465-        7503 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mEGFP-tagged fusion protein, PPP2CA_CDKL3 from 
            patient-derived sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7503)
  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 7503)
  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_PPP2CA_CDKL3
FEATURES             Location/Qualifiers
     source          1..7503
                     /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(2264..2974,2975..2977,2978..2980))
                     /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(2306..2327)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2655..2674
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2914..2935
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2977..2986
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(3062..3265)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3091..3111)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3266..3569
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     CDS             complement(3826..3870)
                     /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    4055..4288
                     /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    4648..4765
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_feature    5257..5382
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             5429..5686
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     enhancer        5688..6067
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6098..6202
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6203..6271,6272..7063)
                     /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(6421..6440)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(7234..7503,1..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 ccggttcacc
     1801 agaagaaaaa gcggttgcgt ttcaggtttt tgccttcgca atgaccatct ccaccttcct
     1861 gtttctcctg gttctggtca acattcaata acgtaggaat tttagaggat tctgttttct
     1921 tagactcacg tttcaacatc ttaatttgtt tgacttccat gcctttcgtg tcttttgact
     1981 ggatggttac cggcaactcc gggaagtgaa actctttacg atccgagcgt gagaaattca
     2041 atttgcgttt attctcgttc gccatctgat cgctatgacc tgtacgctcg ttaaagattc
     2101 ccttctccat ttggcttttt tcgcaaagac tcttgacctg agattcactc agttgtttac
     2161 attcattcaa ctgctcgatc cattgatcca gctctttggt aaaaaccttc tcatccattc
     2221 cccgggaagc ggccgcactt cccccggagc ctccgcttcc gcccttgtac agctcgtcca
     2281 tgccgagagt gatcccggcg gcggtcacga actccagcag gaccatgtga tcgcgcttct
     2341 cgttggggtc tttgctcagc ttggactggg tgctcaggta gtggttgtcg ggcagcagca
     2401 cggggccgtc gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc acgctgccgt
     2461 cctcgatgtt gtggcggatc ttgaagttca ccttgatgcc gttcttctgc ttgtcggcca
     2521 tgatatagac gttgtggctg ttgtagttgt actccagctt gtgccccagg atgttgccgt
     2581 cctccttgaa gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg ccctcgaact
     2641 tcacctcggc gcgggtcttg tagttgccgt cgtccttgaa gaagatggtg cgctcctgga
     2701 cgtagccttc gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc
     2761 ggctgaagca ctgcacgccg taggtcaggg tggtcacgag ggtgggccag ggcacgggca
     2821 gcttgccggt ggtgcagatg aacttcaggg tcagcttgcc gtaggtggca tcgccctcgc
     2881 cctcgccgga cacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg accaggatgg
     2941 gcaccacccc ggtgaacagc tcctcgccct tgctcaccat ggtggcgacc ggtggatccc
     3001 gggcccgcgg taccgtcgac tgcagaattc ggttcactaa acgagcccgg ttcactaaac
     3061 gagctctgct tatatagacc tcccaccgta cacgcctacc gcccatttgc gtcaatgggg
     3121 cggagttgtt acgacatttt ggaaagtccc gttgattttg gtgccaaaac aaactcccat
     3181 tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc aaaccgctat ccacgcccat
     3241 tgatgtactg ccaaaaccgc atcaccatgg taatagcgat gactaatacg tagatgtact
     3301 gccaagtagg aaagtcccat aaggtcatgt actgggcata atgccaggcg ggccatttac
     3361 cgtcattgac gtcaataggg ggcgtacttg gcatatgata cacttgatgt actgccaagt
     3421 gggcagttta ccgtaaatac tccacccatt gacgtcaatg gaaagtccct attggcgtta
     3481 ctatgggaac atacgtcatt attgacgtca atgggcgggg gtcgttgggc ggtcagccag
     3541 gcgggccatt taccgtaagt tatgtaacgc ggaactccat atatgggcta tgaactaatg
     3601 accccgtaat tgattactat taataactag cacgcgttcg aagcttgagc tcggtcccct
     3661 cggggttggg aggtgggtct gaaacgataa tggtgaatat ccctgcctaa ctctattcac
     3721 tatagaaagt acagcaaaaa ctattcttaa acctaccaag cctcctacta tcattatgaa
     3781 taattttata taccacagcc aatttgttat gttaaaccaa ttccacaaac ttgcccattt
     3841 atctaattcc aataattctt gttcattctt ttcttgctgg ttttgcgatt cttcaattaa
     3901 ggagtgtatt aagcttgtgt aattgttaat ttctctgtcc cactccatcc aggtcgtgtg
     3961 attccaaatc tgttccagag atttattact ccaactagca ttccaaggca cagcagtggt
     4021 gcaaatgagt tttccagagc aaccccaaat ccccaggagc tgttgatcct ttaggtatct
     4081 ttccacagcc aggattcttg cctggagctg cttgatgccc cagactgtga gttgcaacag
     4141 atgctgttgc gcctcaatag ccctcagcaa attgttctgc tgctgcacta taccagacaa
     4201 taattgtctg gcctgtaccg tcagcgtcat tgacgctgcg cccatagtgc ttcctgctgc
     4261 tcccaagaac ccaaggaaca aagctcctat tcccactgct cttttttctc tctgcaccac
     4321 tcttctcttt gccttggtgg gtgctactcc taatggttca atttttacta ctttatattt
     4381 atataattca cttctccaat tgtccctcat atctcctcct tatggtgttt tactaaactt
     4441 ttccatgttc taatcctcat cctgtctact tgccacacaa tcatcacctg ccatctgttt
     4501 tccataatcc ctaatgatct ttgcttttct tcttggcact acttttatgt cactattatc
     4561 ttgtattact actgcccctt cacctttcca gaggagcttt gctggtcctt tccaaagtgg
     4621 atttctgctg tccctgtaat aaacccgaaa attttgaatt tttgtaattt gtttttgtaa
     4681 ttctttagtt tgtatgtctg ttgctattat gtctactatt ctttcccctg cactgtaccc
     4741 cccaatcccc ccttttcttt taaaattgtg gatgaatact gccatttgta ctgctgtctt
     4801 aagatgttca gcctgatctc ttacctgtcc tataattttc tttaattctt tattcataga
     4861 ttctactact ccttgacttt ggggattgta gggaatttcc aggtctgaag atcctgctgc
     4921 ttgctgtgct tttttcttac ttttgttttg ctcttcctct atcttgtcta aagcttcctt
     4981 ggtgtctttt atctctatcc tttgatgcac acaatagagg gttgctactg tattatataa
     5041 tgatctaagt tcttctgatc ctgtctgaag ggatggttgt agctgtccca gtatttgtct
     5101 acagccttct gatgtttcta acaggccagt attaactgcg aatcgttcta gctccctgct
     5161 tgcccatact atatgtttta atttatattt tttctttccc cctggcctta accgaatttt
     5221 ttcccatcgc gatctaattc tcccccgctt aatactgacg ctctcgcacc catctctctc
     5281 cttctagcct ccgctagtca aaatttttgg cgtactcacc agtcgccgcc cctcgcctct
     5341 tgccgtgcgc gcttcagcaa gccgagtcct gcgtcgagag agctcctctg gtttcccttt
     5401 cgctttcaag tccctgttcg ggcgccactg ctagagattt tccacactga ctaaaagggt
     5461 ctgagggatc tctagttacc agagtcacac aacagacggg cacacactac ttgaagcact
     5521 caaggcaagc tttattgagg cttaagcagt gggttcccta gttagccaga gagctcccag
     5581 gctcagatct ggtctaacca gagagaccca gtacaggcaa aaagcagctg cttatatgca
     5641 ggatctgagg gctcgccact ccccagtccc gcccaggcca cgcctcccat ggtaatagcg
     5701 atgactaata cgtagatgta ctgccaagta ggaaagtccc ataaggtcat gtactgggca
     5761 taatgccagg cgggccattt accgtcattg acgtcaatag ggggcgtact tggcatatga
     5821 tacacttgat gtactgccaa gtgggcagtt taccgtaaat actccaccca ttgacgtcaa
     5881 tggaaagtcc ctattggcgt tactatggga acatacgtca ttattgacgt caatgggcgg
     5941 gggtcgttgg gcggtcagcc aggcgggcca tttaccgtaa gttatgtaac gcggaactcc
     6001 atatatgggc tatgaactaa tgaccccgta attgattact attaataact agtcaataat
     6061 caatgtcgac ccaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     6121 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     6181 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     6241 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     6301 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     6361 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     6421 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     6481 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     6541 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     6601 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     6661 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     6721 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     6781 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     6841 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     6901 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     6961 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     7021 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     7081 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     7141 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     7201 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     7261 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     7321 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     7381 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     7441 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     7501 cgg
//
