LOCUS       sequence-385204-        7041 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Mammalian Expression of mEGFP-FOXC1 WT Fusion Protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7041)
  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 7041)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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-FOXC1-WT
FEATURES             Location/Qualifiers
     source          1..7041
                     /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      3341..3350
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3354..3488
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3391..3410)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3445..3464
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        3557..3914
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3557..3748
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3580..3600)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3765..3900
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3934..3951)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3934..3950)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3943..3965)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4163..4618
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4250..4269)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4460..4481
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4635..4654)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4754..4776
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4814..4832)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4900..5004
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5005..5073,5074..5865)
                     /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(5223..5242)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6036..6624
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6525..6544
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6778..6795
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6912..6933
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6948..6953,6954..6971,6972..6978)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6986..7002
                     /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     6991..7013
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7010..7026
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7010..7026
                     /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 gcaagctcgg tattcagtta
     1681 gttccccgaa ttctttgggc gtcgttccat atctgggcgg agaacaatct tattatagag
     1741 cagccgcagc tgccgcgggt ggcggatata ctgctatgcc agcgcctatg agtgtttatt
     1801 cccatccagc ccatgcggaa caatatcctg gtggaatggc acgggcttat ggcccatata
     1861 cccctcaacc tcaaccaaaa gatatggtaa aaccaccata ctcttacata gctctgatta
     1921 caatggctat acaaaatgca cccgataaga agattacact taacggtatc taccaattta
     1981 ttatggatag gtttcctttc tacagggata ataaacaagg gtggcaaaat tctattaggc
     2041 ataatctgag tctgaacgaa tgttttgtta aagttccacg ggatgataag aagccaggga
     2101 aaggaagtta ttggactctc gatcccgatt cttataatat gtttgaaaat gggtccttct
     2161 tgcgaaggag aagaaggttt aagaagaaag atgccgtcaa agataaagaa gaaaaggatc
     2221 gattgcatct gaaagaaccc ccaccgcctg gacggcaacc cccacctgcc ccacccgaac
     2281 aagcagatgg taatgcccct ggccctcaac caccaccagt ccggattcaa gatattaaaa
     2341 cggaaaatgg aacatgtccg tctcccccac aaccattgag tccagctgca gctttgggtt
     2401 caggaagtgc tgcggctgta ccaaagatag aatcaccaga tagctccagc tcctcactca
     2461 gctcaggcag ttctccccca ggttccttgc cttctgcaag gcccctgtct ttggatgggg
     2521 ctgactcagc cccaccacca ccagcaccat ctgcaccccc gccccatcac tcccaaggat
     2581 tttcagtcga taatattatg actagtttgc gaggctcacc acaatctgca gcagctgaac
     2641 tgagttcagg tctcttagca agcgctgctg ccagttctcg ggctggaatt gctccgccac
     2701 ttgccttggg cgcttattct cctggacaat cctctcttta ttctagtcct tgttcccaaa
     2761 catcttcagc cggaagttcc ggtggtggtg gtggcggagc tggagcagcc ggcggggcag
     2821 gcggagcggg cacatatcat tgtaatctcc aggcaatgtc attgtatgct gctggagaac
     2881 ggggtggaca tctccaaggg gctcctggcg gtgctggtgg aagcgcagta gatgatcctc
     2941 ttccagatta ttccttaccg cccgtaacga gttcatcaag ctccagcttg agccatggcg
     3001 gtggtggcgg cggtggagga ggcgggcaag aagctgggca tcatcccgca gcacatcagg
     3061 gacggctgac tagctggtat ttaaatcaag ctggcggcga tctcggtcat ctcgcctctg
     3121 ctgctgcagc tgcagccgca gctggatatc ccggacaaca acaaaatttt catagtgtaa
     3181 gagaaatgtt tgaaagccaa cgtattgggc tgaataatag ccctgttaat ggaaacagct
     3241 cttgccagat ggcatttcca agtagtcaaa gtctttatcg aaccagtggt gcctttgttt
     3301 atgattgctc taaattctaa agtcgacctg cagaagcttg gccgccatgg cccaacttgt
     3361 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     3421 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     3481 tctggatcgg gaattaattc ggcgcagcac catggcctga aataacctct gaaagaggaa
     3541 cttggttagg taccttctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt
     3601 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     3661 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3721 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     3781 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     3841 ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     3901 taggcttttg caaaaagctg ttaacagctt ggcactggcc gtcgttttac aacgtcgtga
     3961 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     4021 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     4081 tggcgaatgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     4141 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     4201 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     4261 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     4321 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     4381 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     4441 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     4501 tcgggctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     4561 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     4621 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     4681 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     4741 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     4801 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     4861 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     4921 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     4981 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     5041 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     5101 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     5161 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     5221 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     5281 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     5341 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     5401 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5461 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5521 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     5581 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     5641 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     5701 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     5761 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     5821 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     5881 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     5941 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6001 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     6061 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     6121 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     6181 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     6241 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     6301 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6361 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6421 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6481 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6541 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6601 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6661 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     6721 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     6781 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
     6841 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     6901 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
     6961 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
     7021 tatgacatga ttacgaatta a
//
