LOCUS       sequence-383798-        7757 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of EGFP fused to catalytically dead core p300 histone 
            acetyltransferase (D1399Y).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7757)
  AUTHORS   Schneider MWG, Gibson BA, Otsuka S, Spicer MFD, Petrovic M, Blaukopf
            C, Langer CCH, Batty P, Nagaraju T, Doolittle LK, Rosen MK, Gerlich 
            DW
  TITLE     A mitotic chromatin phase transition prevents perforation by 
            microtubules.
  JOURNAL   Nature. 2022 Sep;609(7925):183-190. doi: 10.1038/s41586-022-05027-y.
            Epub 2022 Aug 3.
  PUBMED    35922507
REFERENCE   2  (bases 1 to 7757)
  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/s41586-022-05027-y"; journalName: "Nature"; date: 
            "2022-09"; volume: "609"; issue: "7925"; pages: "183-190"
COMMENT     Sequence Label: EGFP-p300 (D1399Y)
FEATURES             Location/Qualifiers
     source          1..7757
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        87..191
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(192..260,261..1052)
                     /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(410..429)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1223..1811
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1712..1731
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1965..1982
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2099..2120
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2135..2140,2141..2158,2159..2165)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2173..2189
                     /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     2178..2200
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2197..2213
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2197..2213
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2273..2292)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    2339..2563
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2552..2569
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(2705..3304)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2788..2808)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     3285..3304
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     misc_feature    join(3324..3332,3333..3335,3336..3910)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3497..3516)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     3726..3743
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     complement(3980..3999)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          3982..4211
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     CDS             complement(4342..4368)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(4369..6219)
                     /codon_start=1
                     /gene="EP300"
                     /product="catalytic histone acetyltransferase core of the 
                     human E1A-associated protein p300, inactivated by the 
                     D1399Y point mutation (Hilton et al., 2015)"
                     /label=p300 core (D1399Y)
                     /translation="IFKPEELRQALMPTLEALYRQDPESLPFRQPVDPQLLGIPDYFDI
                     VKSPMDLSTIKRKLDTGQYQEPWQYVDDIWLMFNNAWLYNRKTSRVYKYCSKLSEVFEQ
                     EIDPVMQSLGYCCGRKLEFSPQTLCCYGKQLCTIPRDATYYSYQNRYHFCEKCFNEIQG
                     ESVSLGDDPSQPQTTINKEQFSKRKNDTLDPELFVECTECGRKMHQICVLHHEIIWPAG
                     FVCDGCLKKSARTRKENKFSAKRLPSTRLGTFLENRVNDFLRRQNHPESGEVTVRVVHA
                     SDKTVEVKPGMKARFVDSGEMAESFPYRTKALFAFEEIDGVDLCFFGMHVQEYGSDCPP
                     PNQRRVYISYLYSVHFFRPKCLRTAVYHEILIGYLEYVKKLGYTTGHIWACPPSEGDDY
                     IFHCHPPDQKIPKPKRLQEWYKKMLDKAVSERIVHDYKDIFKQATEDRLTSAKELPYFE
                     GDFWPNVLEESIKELEQEEEERKREENTSNESTDVTKGDSKNAKKKNNKKTSKNKSSLS
                     RGNKKKPGMPNVSNDLSQKLYATMEKHKEVFFVIRLIAGPAANSLPPIVDPDPLIPCDL
                     MDGRDAFLTLARDKHLEFSSLRRAQWSTMCMLVELHTQSQD"
     regulatory      4596..4605
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     6644..6663
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             complement(6994..7263)
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="EMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTLKETS
                     FNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISKQ"
     CDS             complement(6997..7275)
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
     promoter        complement(7298..7509)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     primer_bind     join(7750..7757,1..12)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gagcagattg tactgagagt gcaccatagg ggatcgggag atctcccgat ccgtcgacgt
       61 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
      121 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
      181 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
      241 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
      301 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
      361 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
      421 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
      481 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
      541 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
      601 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
      661 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
      721 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
      781 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
      841 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
      901 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
      961 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     1021 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     1081 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     1141 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     1201 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     1261 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     1321 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     1381 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     1441 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     1501 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     1561 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     1621 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     1681 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     1741 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     1801 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     1861 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     1921 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     1981 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     2041 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     2101 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     2161 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
     2221 acgccaagct ctagctagag gtcgacggta tacagacatg ataagataca ttgatgagtt
     2281 tggacaaacc acaactagaa tgcactcgag ccccagctgg ttctttccgc ctcagaagcc
     2341 atagagccca ccgcatcccc agcatgcctg ctattgtctt cccaatcctc ccccttgctg
     2401 tcctgcccca ccccaccccc cagaatagaa tgacacctac tcagacaatg cgatgcaatt
     2461 tcctcatttt attaggaaag gacagtggga gtggcacctt ccagggtcaa ggaaggcacg
     2521 ggggaggggc aaacaacaga tggctggcaa ctagaaggca cagtcgaggc tgatcagcga
     2581 gctctagagt cggtgggcct cgggggcggg tgcggggtcg gcggggccgc cccgggtggc
     2641 ttcggtcgga gccatggggt cgtgcgctcc tttcggtcgg gcgctgcggg tcgtggggcg
     2701 ggcgtcaggc accgggcttg cgggtcatgc accaggtgcg cggtccttcg ggcacctcga
     2761 cgtcggcggt gacggtgaag ccgagccgct cgtagaaggg gaggttgcgg ggcgcggagg
     2821 tctccaggaa ggcgggcacc ccggcgcgct cggccgcctc cactccgggg agcacgacgg
     2881 cgctgcccag acccttgccc tggtggtcgg gcgagacgcc gacggtggcc aggaaccacg
     2941 cgggctcctt gggccggtgc ggcgccagga ggccttccat ctgttgctgc gcggccagcc
     3001 gggaaccgct caactcggcc atgcgcgggc cgatctcggc gaacaccgcc cccgcttcga
     3061 cgctctccgg cgtggtccag accgccaccg cggcgccgtc gtccgcgacc cacaccttgc
     3121 cgatgtcgag cccgacgcgc gtgaggaaga gttcttgcag ctcggtgacc cgctcgatgt
     3181 ggcggtccgg gtcgacggtg tggcgcgtgg cggggtagtc ggcgaacgcg gcggcgaggg
     3241 tgcgtacggc ccgggggacg tcgtcgcggg tggcgaggcg caccgtgggc ttgtactcgg
     3301 tcatggaagg tcgtctcctt gtgggttgtg gcaagcttat catcgtgttt ttcaaaggaa
     3361 aaccacgtcc ccgtggttcg gggggcctag acgttttttt aacctcgact aaacacatgt
     3421 aaagcatgtg caccgaggcc ccagatcaga tcccatacaa tggggtacct tctgggcatc
     3481 cttcagcccc ttgttgaata cgcttgagga gagccatttg actctttcca caactatcca
     3541 actcacaacg tggcactggg gttgtgccgc ctttgcaggt gtatcttata cacgtggctt
     3601 ttggccgcag aggcacctgt cgccaggtgg ggggttccgc tgcctgcaaa gggtcgctac
     3661 agacgttgtt tgtcttcaag aagcttccag aggaactgct tccttcacga cattcaacag
     3721 accttgcatt cctttggcga gaggggaaag acccctagga atgctcgtca agaagacagg
     3781 gccaggtttc cgggccctca cattgccaaa agacggcaat atggtggaaa ataacatata
     3841 gacaaacgca caccggcctt attccaagcg gcttcggcca gtaacgttag ggggggggga
     3901 gggagagggg gggggggaga ggggcggaat tggccgccct agatgcatgc tcgacctgca
     3961 gttggacctg ggagtggaca cctgtggaga gaaaggcaaa gtggatgtca ttgtcactca
     4021 agtgtatggc cagatctcaa gcctgccaca cctcaagctt gacaacaaaa agattgtctt
     4081 ttctgaccag atggacgcgg ccaccctcaa aggcatcacc gcgggccagg tgaatatcaa
     4141 atcctcctcg tttttggaaa ctgacaatct tagcgcagaa gtcatgcccg cttttgagag
     4201 ggagtactca ccccaacagc tggccctcgc agacagcgaa ttaattccag cacactggat
     4261 cagttatcta tgcggccgcg agctcggatc cactagtaac ggccgccagt gtgctggaat
     4321 tctgcagata attcgtcaag aagcgtagtc cggaacgtcg tacgggtagt cctggctctg
     4381 cgtgtgcagc tccaccagca tgcacatggt ggaccactgg gctcttcgga gtgaagagaa
     4441 ctccaggtgc ttgtcccttg ccagcgtgag aaacgcatcc cgaccatcca tcagatcgca
     4501 ggggatgaga ggatcaggat caacaatggg aggcagggag ttggcagcag ggccagcaat
     4561 gaggcggatc acaaagaaga cctctttatg cttctccatg gtggcatata gtttctgtga
     4621 gaggtcgtta gatacattgg gcatcccggg tttcttcttg ttgcccctac tcaggctgct
     4681 cttatttttg ctggttttct tattattctt ctttttagca tttttgctgt ctcccttggt
     4741 cacatctgtg ctttcattgc tggtgttttc ctctcgtttt ctctcttctt cctcctgttc
     4801 cagttcctta atgctttctt ccagaacatt gggccagaaa tcaccctcga aataaggcaa
     4861 ttcctttgca cttgttaatc tatcttcagt agcttgttta aaaatatcct tgtagtcatg
     4921 gacaatacgc tctgatacag ccttgtcaag catttttttg taccattcct gcagtcgctt
     4981 gggcttgggt atcttctggt caggaggatg gcaatggaag atataatcat ctccctcact
     5041 tggtggacat gcccaaatat gccctgttgt gtaacctaat ttcttgacat attctaaata
     5101 tccaattagg atttcatgat agactgcagt cctcaagcat ttaggacgga agaaatgaac
     5161 actatagagg taagatatgt atactctcct ctggttgggt ggagggcagt cagagccata
     5221 ctcttgaaca tgcatgccaa agaagcacag gtcaacacca tcaatttctt caaaggcaaa
     5281 gagggctttg gttcggtatg gaaaggattc tgccatctct ccactgtcca caaaccttgc
     5341 tttcatgcct ggttttactt ccacggtttt gtcagaagca tgaactactc taacagtgac
     5401 ctctcctgac tcagggtgat tctgtcgcct cagaaagtca ttcacacgat tctctagaaa
     5461 ggtgccaagt ctggtagatg gcaacctttt agcagaaaac ttattttctt tcctagttcg
     5521 tgcacttttc tttaaacagc catcacagac gaatccagca ggccagatga tctcatggtg
     5581 aaggacacag atctgatgca tctttcttcc gcactctgta cattcaacaa acagttcagg
     5641 atccagtgtg tcattttttc tcttggaaaa ttgttcttta tttattgtag tttgaggctg
     5701 ggaagggtca tcccccaaag aaacgctctc cccttggatc tcattgaaac acttctcaca
     5761 gaaatgatac ctgttctggt aactgtaata agtggcatca cgaggtattg tgcacaactg
     5821 tttgccgtag caacacagtg tctgtggaga gaactccaac tttctgccac aacagtatcc
     5881 aaggctttgc atcactgggt caatttcttg ttcaaagacc tcagagagct tggagcagta
     5941 tttgtatacc cgtgatgttt tccggttata taaccaggca ttattgaaca taagccaaat
     6001 atcatcgaca tactgccagg gctcctgata ctgtccagtg tctaacttcc tcttaatggt
     6061 agaaagatcc atggggctct tcacaatatc aaagtaatca gggattccta aaagctgagg
     6121 gtccacaggt tgacgaaagg gaagggattc tggatcctga cggtaaagtg cctccaaagt
     6181 tggcatcagt gcctgtcgta gttcttctgg tttgaaaatg gcgcgcccgc tagccttgga
     6241 tccggcaatt ggcttgtaca gctcgtccat gccgagagtg atcccggctg cggtcacgaa
     6301 ctccttaagg accatgtgat ctcgcttctc gttggggtct ttgctcaggg cggactgggt
     6361 gctcaggtag tggttgtcag gcagcagcac gggtccgtca ccgattgggg tgttctgctg
     6421 gtagtggtcg gcgagctgca cgcttccgtc ctcgatgttg tgacggatct tgaagttcac
     6481 cttgattccg ttcttctgct tgtcagccat gatatagacg ttgtggctgt tgtagttgta
     6541 ctccagcttg tgacccagga tgttaccgtc ctccttgaag tcgatgccct tcagctcgat
     6601 gcggttcacc agggtgtctc cctcgaactt cacctcagcg cgagtcttgt agttgccgtc
     6661 gtccttgaag aagatggtac gctcctggac gtagccttcg ggcatggcgg acttgaagaa
     6721 gtcgtgctgc ttcatgtggt cggggtatcg gctgaagcac tgcacgccgt aggtcagggt
     6781 ggtcacgagg gtaggccagg gcacgggcag cttgccggtg gtgcagatga acttcagggt
     6841 cagcttgccg taggtagcat cgccctctcc ctcaccactc acactgaact tgtgaccgtt
     6901 tacgtcaccg tccagttcga ccaggattgg caccactccg gtgaacagct cctctccctt
     6961 gctcacggat cctccacctc cacctccgcc ggcctgcttt gagattcgtc ggaacacatg
     7021 ataatagagg tcccaggctt gggtgaggtc cttgacattc cctgatttca tgtacttcct
     7081 gcaccactct tgggcctcca ttaaatctcg accataggcc tgattaaagg atgtttcctt
     7141 cagagtctgg gggccccgtt ccatcatagc atgcaagggc tccagcacct caaacatgcc
     7201 tttcacgttc ctttccccaa agtacaaacg agatgcctct tccaggcctt catgccacat
     7261 ctcatgccag aggatcatgg tggctctaga accggtcctg tgttctggcg gcaaacccgt
     7321 tgcgaaaaag aacgttcacg gcgactactg cacttatata cggttctccc ccaccctcgg
     7381 gaaaaaggcg gagccagtac acgacatcac tttcccagtt taccccgcgc caccttctct
     7441 aggcaccgga tcaattgccg acccctcccc ccaacttctc ggggactgtg ggcgatgtgc
     7501 gctctgccca ctgacgggca ccggagccaa ttcccactcc tttcaagacc tagctagcac
     7561 tagtcaataa tcaatgtcaa cgcgtatatc tggcccgtac atcgcgaagc agcgcaaaac
     7621 gcctaaccct aagcagattc ttcatgcaat tgtcggtcaa gccttgcctt gttgtagctt
     7681 aaattttgct cgcgcactac tcagcgacct ccaacacaca agcagggagc agatactggc
     7741 ttaactatgc ggcatca
//
