LOCUS       sequence-385244-        7311 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of mEGFP-DVL1 Mutant (H502Pfs*143) Fusion 
            Protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7311)
  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 7311)
  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-DVL1-MUT
FEATURES             Location/Qualifiers
     source          1..7311
                     /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      3611..3620
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3624..3758
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3661..3680)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3715..3734
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        3827..4184
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3827..4018
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3850..3870)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4035..4170
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4204..4221)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4204..4220)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4213..4235)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4433..4888
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4520..4539)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4730..4751
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4905..4924)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5024..5046
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5084..5102)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5170..5274
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5275..5343,5344..6135)
                     /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(5493..5512)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6306..6894
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6795..6814
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7048..7065
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7182..7203
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7218..7223,7224..7241,7242..7248)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7256..7272
                     /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     7261..7283
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7280..7296
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7280..7296
                     /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 ggcggagacc aagattatct
     1681 accacatgga cgaggaggag acgccgtacc tggtcaagct gcccgtggcc cccgagcgcg
     1741 tcacgctggc cgacttcaag aacgtgctca gcaaccggcc cgtgcacgcc tacaaattct
     1801 tctttaagtc catggaccag gacttcgggg tggtgaagga ggagatcttt gatgacaatg
     1861 ccaagcttcc ctgcttcaac ggccgcgtgg tctcctggct ggtcctggct gagggtgctc
     1921 actcggatgc ggggtcccag ggcacggaca gccacacaga cctgcccccg cctcttgagc
     1981 ggacaggcgg catcggggac tcccggcccc cctccttcca cccaaatgtg gccagcagcc
     2041 gtgacgggat ggacaacgag acaggcacgg agtccatggt cagtcaccgg cgggagcgtg
     2101 cccgacgccg gaaccgcgag gaggccgccc ggaccaatgg gcacccaagg ggagaccgac
     2161 ggcgggatgt ggggctgccc ccagacagcg cgtccaccgc cctcagcagc gagcttgagt
     2221 ccagcagctt tgtggactcg gacgaggatg gcagcacgag caggctcagc agctccacgg
     2281 agcagagcac ctcatccaga ctcatccgga agcacaaacg ccggcggagg aagcagcgcc
     2341 ttcggcaggc ggaccgggcc tcctccttca gcagcataac cgactccacc atgtccctca
     2401 acatcgtcac tgtcacgctc aacatggaaa gacatcactt tctgggcatc agcatcgtgg
     2461 ggcagagcaa cgaccgtgga gacggcggca tctacattgg ctccatcatg aagggcgggg
     2521 ctgtggccgc tgacggccgc atcgagcccg gcgacatgtt gctgcaggtg aatgacgtga
     2581 actttgagaa catgagcaat gacgatgccg tgcgggtgct gcgggagatc gtttcccaga
     2641 cggggcccat cagcctcact gtggccaagt gctgggaccc aacgccccga agctacttca
     2701 ccgtcccacg ggctgacccg gtgcggccca tcgaccccgc cgcctggctg tcccacacgg
     2761 cggcactgac aggagccctg ccccgctacg agctggaaga ggcgccgctg acggtgaaga
     2821 gtgacatgag cgccgtcgtc cgggtcatgc aattgcccga ttccgggctt gaaatacgag
     2881 ataggatgtg gctgaaaatt actattgcta acgcggtgat tggcgccgat gtcgtcgatt
     2941 ggctctatac tcatgtcgaa gggtttaaag aacgcagaga agcaagaaaa tatgcgtcct
     3001 ctctgctcaa acatgggttt ctcagacata ccgtgaataa aattactttc tctgaacaat
     3061 gttattatgt gtttggagac ctgtgtagta acctggcaac gctcaatctg aattccggaa
     3121 gtagcggtac atctgaccaa gatactcttg cacctcttcc accagggctt tgggtgcggg
     3181 cgacaccaac atcaacacgc gatccacatc ccgcatctcg gctcccaact cgtactcgtg
     3241 ccctggcaat ggccgctgcc gctccagggg ttagtagggt aaaaggcgcc aaggctgttg
     3301 gaccaccagg cgctgctgct ggacctcgag ctgtccggag atccgttgga aggagagaac
     3361 tcggagccgt cgccgtcaac cgtataaccc gacatagagt tggctggggt gctgcgggag
     3421 aatctgttcg cccagcatca tctgcagtcg ccgccgctca tgctgtgagg ccaagacttc
     3481 cccctagagg gagcccacgg ccaacacctc ggcccagacc catccaatgg tggggtggac
     3541 atccagggga ccccctgtcc gggagctggc tgccgtcccc ccggaattga agtcgacctg
     3601 cagaagcttg gccgccatgg cccaacttgt ttattgcagc ttataatggt tacaaataaa
     3661 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     3721 tgtccaaact catcaatgta tcttatcatg tctggatcgg gaattaattc ggcgcagcac
     3781 catggcctga aataacctct gaaagaggaa cttggttagg taccttctga ggcggaaaga
     3841 accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     3901 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     3961 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     4021 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     4081 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     4141 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctg ttaacagctt
     4201 ggcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
     4261 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
     4321 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct
     4381 ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc
     4441 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     4501 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     4561 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     4621 cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc
     4681 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     4741 ttccaaactg gaacaacact caaccctatc tcgggctatt cttttgattt ataagggatt
     4801 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     4861 tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat
     4921 gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     4981 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     5041 cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta
     5101 tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg
     5161 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     5221 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     5281 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     5341 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     5401 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     5461 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     5521 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     5581 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     5641 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     5701 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     5761 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     5821 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     5881 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     5941 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     6001 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     6061 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     6121 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     6181 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     6241 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     6301 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     6361 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     6421 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     6481 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     6541 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     6601 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     6661 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     6721 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     6781 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     6841 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     6901 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     6961 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     7021 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     7081 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     7141 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact
     7201 cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg
     7261 agcggataac aatttcacac aggaaacagc tatgacatga ttacgaatta a
//
