LOCUS       sequence-436097-        8133 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of an EZH2 catalytic site mutant in insect cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8133)
  AUTHORS   Gail EH, Healy E, Flanigan SF, Jones N, Ng XH, Uckelmann M, Levina 
            V, Zhang Q, Davidovich C
  TITLE     Inseparable RNA binding and chromatin modification activities of a 
            nucleosome-interacting surface in EZH2.
  JOURNAL   Nat Genet. 2024 May 14. doi: 10.1038/s41588-024-01740-8.
  PUBMED    38744974
REFERENCE   2  (bases 1 to 8133)
  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 Genet. 
            2024 May 14. doi: 10.1038/s41588-024-01740-8."
COMMENT     Sequence Label: pFB1.HMBP.PrS.dEZH2
FEATURES             Location/Qualifiers
     source          1..8133
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     732..749
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  883..1107
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(1174..1707)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(1877..1882,1883..1899,1900..1905))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     promoter        2257..2348
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     2273..2290
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             2409..2426
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2433..3530
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="KTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE
                     KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI
                     AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA
                     DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET
                     AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE
                     NYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFW
                     YAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      3387..3396
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     3504..3527
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             3558..3581
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     polyA_signal    6006..6140
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6043..6062)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6097..6116
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  6169..6334
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      6518..6972
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6605..6624)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6814..6835
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6998..7102
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7103..7171,7172..7963)
                     /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(7321..7340)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
      661 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
      721 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc
      781 tccttacgca tctgtgcggt atttcacacc gcatagacca gccgcgtaac ctggcaaaat
      841 cggttacggt tgagtaataa atggatgccc tgcgtaagcg ggtgtgggcg gacaataaag
      901 tcttaaactg aacaaaatag atctaaacta tgacaataaa gtcttaaact agacagaata
      961 gttgtaaact gaaatcagtc cagttatgct gtgaaaaagc atactggact tttgttatgg
     1021 ctaaagcaaa ctcttcattt tctgaagtgc aaattgcccg tcgtattaaa gaggggcgtg
     1081 gccaagggca tggtaaagac tatattcgcg gcgttgtgac aatttaccga acaactccgc
     1141 ggccgggaag ccgatctcgg cttgaacgaa ttgttaggtg gcggtacttg ggtcgatatc
     1201 aaagtgcatc acttcttccc gtatgcccaa ctttgtatag agagccactg cgggatcgtc
     1261 accgtaatct gcttgcacgt agatcacata agcaccaagc gcgttggcct catgcttgag
     1321 gagattgatg agcgcggtgg caatgccctg cctccggtgc tcgccggaga ctgcgagatc
     1381 atagatatag atctcactac gcggctgctc aaacttgggc agaacgtaag ccgcgagagc
     1441 gccaacaacc gcttcttggt cgaaggcagc aagcgcgatg aatgtcttac tacggagcaa
     1501 gttcccgagg taatcggagt ccggctgatg ttgggagtag gtggctacgt ctccgaactc
     1561 acgaccgaaa agatcaagag cagcccgcat ggatttgact tggtcagggc cgagcctaca
     1621 tgtgcgaatg atgcccatac ttgagccacc taactttgtt ttagggcgac tgccctgctg
     1681 cgtaacatcg ttgctgctgc gtaacatcaa acatcgaccc acggcgtaac gcgcttgctg
     1741 cttggatgcc cgaggcatag actgtacaaa aaaacagtca taacaagcca tgaaaaccgc
     1801 cactgcgccg ttaccaccgc tgcgttcggt caaggttctg gaccagttgc gtgagcgcat
     1861 acgctacttg cattacagtt tacgaaccga acaggcttat gtcaactggg ttcgtgcctt
     1921 catccgtttc cacggtgtgc gtcacccggc aaccttgggc agcagcgaag tcgaggcatt
     1981 tctgtcctgg ctggcgaacg agcgcaaggt ttcggtctcc acgcatcgtc aggcattggc
     2041 ggccttgctg ttcttctacg gcaaggtgct gtgcacggat ctgccctggc ttcaggagat
     2101 cggaagacct cggccgtcgc ggcgcttgcc ggtggtgctg accccggatg aagtggttcg
     2161 catcctcggt tttctggaag gcgagcatcg tttgttcgcc caggactcta gctatagttc
     2221 tagtggttgg ctacgtatac tccggaatat taatagatca tggagataat taaaatgata
     2281 accatctcgc aaataaataa gtattttact gttttcgtaa cagttttgta ataaaaaaac
     2341 ctataaatat tccggattat tcataccgtc ccaccatcgg gcgcggatct cggtccgaaa
     2401 ccatgggtca tcaccatcat catcacggta ccaaaactga agaaggtaaa ctggtaatct
     2461 ggattaacgg cgataaaggc tataacggtc tcgctgaagt cggtaagaaa ttcgagaaag
     2521 ataccggaat taaagtcacc gttgagcatc cggataaact ggaagagaaa ttcccacagg
     2581 ttgcggcaac tggcgatggc cctgacatta tcttctgggc acacgaccgc tttggtggct
     2641 acgctcaatc tggcctgttg gctgaaatca ccccggacaa agcgttccag gacaagctgt
     2701 atccgtttac ctgggatgcc gtacgttaca acggcaagct gattgcttac ccgatcgctg
     2761 ttgaagcgtt atcgctgatt tataacaaag atctgctgcc gaacccgcca aaaacctggg
     2821 aagagatccc ggcgctggat aaagaactga aagcgaaagg taagagcgcg ctgatgttca
     2881 acctgcaaga accgtacttc acctggccgc tgattgctgc tgacgggggt tatgcgttca
     2941 agtatgaaaa cggcaagtac gacattaaag acgtgggcgt ggataacgct ggcgcgaaag
     3001 cgggtctgac cttcctggtt gacctgatta aaaacaaaca catgaatgca gacaccgatt
     3061 actccatcgc agaagctgcc tttaataaag gcgaaacagc gatgaccatc aacggcccgt
     3121 gggcatggtc caacatcgac accagcaaag tgaattatgg tgtaacggta ctgccgacct
     3181 tcaagggtca accatccaaa ccgttcgttg gcgtgctgag cgcaggtatt aacgccgcca
     3241 gtccgaacaa agagctggca aaagagttcc tcgaaaacta tctgctgact gatgaaggtc
     3301 tggaagcggt taataaagac aaaccgctgg gtgccgtagc gctgaagtct tacgaggaag
     3361 agttggcgaa agatccacgt attgccgcca ccatggaaaa cgcccagaaa ggtgaaatca
     3421 tgccgaacat cccgcagatg tccgctttct ggtatgccgt gcgtactgcg gtgatcaacg
     3481 ccgccagcgg tcgtcagact gtcgatgaag ccctgaaaga cgcgcagact ggtaccgatt
     3541 acgatatccc aacgaccctg gaagttctgt tccaggggcc cctgggatcc ggccagactg
     3601 ggaagaaatc tgagaaggga ccagtttgtt ggcggaagcg tgtaaaatca gagtacatgc
     3661 gactgagaca gctcaagagg ttcagacgag ctgatgaagt aaagagtatg tttagttcca
     3721 atcgtcagaa aattttggaa agaacggaaa tcttaaacca agaatggaaa cagcgaagga
     3781 tacagcctgt gcacatcctg acttctgtga gctcattgcg cgggactagg gagtgttcgg
     3841 tgaccagtga cttggatttt ccaacacaag tcatcccatt aaagactctg aatgcagttg
     3901 cttcagtacc cataatgtat tcttggtcac ccctacagca gaattttatg gtggaagatg
     3961 aaactgtttt acataacatt ccttatatgg gagatgaagt tttagatcag gatggtactt
     4021 tcattgaaga actaataaaa aattatgatg ggaaagtaca cggggataga gaatgtgggt
     4081 ttataaatga tgaaattttt gtggagttgg tgaatgccct tggtcaatat aatgatgatg
     4141 acgatgatga tgatggagac gatcctgaag aaagagaaga aaagcagaaa gatctggagg
     4201 atcaccgaga tgataaagaa agccgcccac ctcggaaatt tccttctgat aaaatttttg
     4261 aagccatttc ctcaatgttt ccagataagg gcacagcaga agaactaaag gaaaaatata
     4321 aagaactcac cgaacagcag ctcccaggcg cacttcctcc tgaatgtacc cccaacatag
     4381 atggaccaaa tgctaaatct gttcagagag agcaaagctt acactccttt catacgcttt
     4441 tctgtaggcg atgttttaaa tatgactgct tcctacatcg taagtgcaat tattcttttc
     4501 atgcaacacc caacacttat aagcggaaga acacagaaac agctctagac aacaaacctt
     4561 gtggaccaca gtgttaccag catttggagg gagcaaagga gtttgctgct gctctcaccg
     4621 ctgagcggat aaagacccca ccaaaacgtc caggaggccg cagaagagga cggcttccca
     4681 ataacagtag caggcccagc acccccacca ttaatgtgct ggaatcaaag gatacagaca
     4741 gtgataggga agcagggact gaaacggggg gagagaacaa tgataaagaa gaagaagaga
     4801 agaaagatga aacttcgagc tcctctgaag caaattctcg gtgtcaaaca ccaataaaga
     4861 tgaagccaaa tattgaacct cctgagaatg tggagtggag tggtgctgaa gcctcaatgt
     4921 ttagagtcct cattggcact tactatgaca atttctgtgc cattgctagg ttaattggga
     4981 ccaaaacatg tagacaggtg tatgagttta gagtcaaaga atctagcatc atagctccag
     5041 ctcccgctga ggatgtggat actcctccaa ggaaaaagaa gaggaaacac cggttgtggg
     5101 ctgcacactg cagaaagata cagctgaaaa aggacggctc ctctaaccat gtttacaact
     5161 atcaaccctg tgatcatcca cggcagcctt gtgacagttc gtgcccttgt gtgatagcac
     5221 aaaatttttg tgaaaagttt tgtcaatgta gttcagagtg tcaaaaccgc tttccgggat
     5281 gccgctgcaa agcacagtgc aacaccaagc agtgcccgtg ctacctggct gtccgagagt
     5341 gtgaccctga cctctgtctt acttgtggag ccgctgacca ttgggacagt aaaaatgtgt
     5401 cctgcaagaa ctgcagtatt cagcggggct ccaaaaagca tctattgctg gcaccatctg
     5461 acgtggcagg ctgggggatt tttatcaaag atcctgtgca gaaaaatgaa ttcatctcag
     5521 aatactgtgg agagattatt tctcaagatg aagctgacag aagagggaaa gtgtatgata
     5581 aatacatgtg cagctttctg ttcaacttga acaatgattt tgtggtggat gcaacccgca
     5641 agggtaacaa aattcgtttt gcaaatcatt cggtaaatcc aaactgctat gcaaaagtta
     5701 tgatggttaa cggtgatcac aggataggta tttttgccaa gagagccatc cagactggcg
     5761 aagagctgtt ttttgatttc agatacagcc aggctgatgc cctgaagtat gtcggcatcg
     5821 aaagagaaat ggaaatccct tgatctagag cctgcagtct cgaggcatgc ggtaccaagc
     5881 ttgtcgagaa gtactagagg atcataatca gccataccac atttgtagag gttttacttg
     5941 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     6001 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     6061 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     6121 tatcttatca tgtctggatc tgatcactga tatcgcctag gagatccgaa ccagataagt
     6181 gaaatctagt tccaaactat tttgtcattt ttaattttcg tattagctta cgacgctaca
     6241 cccagttccc atctattttg tcactcttcc ctaaataatc cttaaaaact ccatttccac
     6301 ccctcccagt tcccaactat tttgtccgcc cacagcgggg catttttctt cctgttatgt
     6361 ttttaatcaa acatcctgcc aactccatgt gacaaaccgt catcttcggc tactttttct
     6421 ctgtcacaga atgaaaattt ttctgtcatc tcttcgttat taatgtttgt aattgactga
     6481 atatcaacgc ttatttgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta
     6541 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     6601 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     6661 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     6721 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     6781 cgccctttga cgttggagtc cacgttctta atagtggact cttgttccaa actggaacaa
     6841 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     6901 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     6961 cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     7021 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     7081 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     7141 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     7201 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     7261 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     7321 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     7381 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     7441 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     7501 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     7561 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     7621 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     7681 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     7741 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     7801 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     7861 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     7921 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     7981 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     8041 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     8101 gtcagacccc gtagaaaaga tcaaaggatc ttc
//
