LOCUS       sequence-248506-        6767 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses human EZH2 in insect cells, under a TEV-cleavable 
            N-terminal polyhistidine tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 6767)
  AUTHORS   Zhang Q, McKenzie NJ, Warneford-Thomson R, Gail EH, Flanigan SF, 
            Owen BM, Lauman R, Levina V, Garcia BA, Schittenhelm RB, Bonasio R, 
            Davidovich C
  TITLE     RNA exploits an exposed regulatory site to inhibit the enzymatic 
            activity of PRC2.
  JOURNAL   Nat Struct Mol Biol. 2019 Mar;26(3):237-247. doi: 
            10.1038/s41594-019-0197-y. Epub 2019 Mar 4.
  PUBMED    30833789
REFERENCE   2  (bases 1 to 6767)
  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/s41594-019-0197-y"; journalName: "Nat Struct Mol Biol"; 
            date: "2019-03"; volume: "26"; issue: "3"; pages: "237-247"
COMMENT     Sequence Label: pFBOH-MHL.EZH2
FEATURES             Location/Qualifiers
     source          1..6767
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     rep_origin      2..457
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(89..108)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     299..320
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        484..588
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(589..657,658..1449)
                     /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(807..826)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1620..2208
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2109..2128
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2362..2379
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  2511..2735
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(2802..3335)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPRFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3524..3529,3530..3546,3547..3552))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     promoter        3904..3995
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     3920..3937
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             4054..4071
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4084..4104
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     protein_bind    complement(4132..4165)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        4180..4625
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(4626..4712,4713..6047)
                     /codon_start=1
                     /gene="Bacillus subtilis sacB"
                     /product="secreted levansucrase that renders bacterial 
                     growth sensitive to sucrose"
                     /label=SacB
                     /note="negative selection marker"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYKVPEFDSSTIKNISSAKGLDVWDSWPLQNTDGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     polyA_signal    6257..6391
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6294..6313)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6348..6367
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  6420..6585
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
       61 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
      121 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
      181 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
      241 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
      301 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
      361 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
      421 aacgcgaatt ttaacaaaat attaacgttt acaatttcag gtggcacttt tcggggaaat
      481 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
      541 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
      601 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
      661 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
      721 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
      781 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
      841 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
      901 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
      961 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     1021 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     1081 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     1141 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     1201 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     1261 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     1321 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     1381 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     1441 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     1501 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     1561 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     1621 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     1681 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     1741 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     1801 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     1861 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     1921 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     1981 tacaccgaac tgagatacct acagcgtgag cattgagaaa gcgccacgct tcccgaaggg
     2041 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     2101 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     2161 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     2221 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     2281 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     2341 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     2401 cggtattttc tccttacgca tctgtgcggt atttcacacc gcagaccagc cgcgtaacct
     2461 ggcaaaatcg gttacggttg agtaataaat ggatgccctg cgtaagcggg tgtgggcgga
     2521 caataaagtc ttaaactgaa caaaatagat ctaaactatg acaataaagt cttaaactag
     2581 acagaatagt tgtaaactga aatcagtcca gttatgctgt gaaaaagcat actggacttt
     2641 tgttatggct aaagcaaact cttcattttc tgaagtgcaa attgcccgtc gtattaaaga
     2701 ggggcgtggc caagggcatg gtaaagacta tattcgcggc gttgtgacaa tttaccgaac
     2761 aactccgcgg ccgggaagcc gatctcggct tgaacgaatt gttaggtggc ggtacttggg
     2821 tcgatatcaa agtgcatcac ttcttcccgt atgcccaact ttgtatagag agccactgcg
     2881 ggatcgtcac cgtaatctgc ttgcacgtag atcacataag caccaagcgc gttggcctca
     2941 tgcttgagca gattgatgag cgcggtggca atgccctgcc tccggtgctc gccggagact
     3001 gcgagatcat agatatagat ctcactacgc ggctgctcaa acctgggcag aacgtaagcc
     3061 gcgagagcgc caacaaccgc ttcttggtcg aaggcagcaa gcgcgatgaa tgtcttacta
     3121 cggagcaagt tcccgaggta atcggagtcc ggctgatgtt gggagtaggt ggctacgtct
     3181 ccgaactcac gaccgaaaag atcaagagca gcccgcatgg atttgacttg gtcagggccg
     3241 agcctacatg tgcgaatgat gcccatactt gagccaccta actttgtttt agggcgactg
     3301 ccctgctgcg taacatcgtt gctgctgcgt aacatcgttg ctgctccata acatcaaaca
     3361 tcgacccacg gcgtaacgcg cttgctgctt ggatgcccga ggcatagact gtacaaaaaa
     3421 acagtcataa caagccatga aaaccgccac tgcgccgtta ccaccgctgc gttcggtcaa
     3481 ggttctggac cagttgcgtg agcgcatacg ctacttgcat tacagtttac gaaccgaaca
     3541 ggcttatgtc aactgggttc gtgccttcat ccgtttccac ggtgtgcgtc acccggcaac
     3601 cttgggcagc agcgaagtcg aggcatttct gtcctggctg gcgaacgagc gcaaggtttc
     3661 ggtctccacg catcgtcagg cattggcggc cttgctgttc ttctacggca aggtgctgtg
     3721 cacggatctg ccctggcttc aggagatcgg aagacctcgg ccgtcgcggc gcttgccggt
     3781 ggtgctgacc ccggatgaag tggttcgcat cctcggtttt ctggaaggcg agcatcgttt
     3841 gttcgcccag gactctagct atagttctag tggttggcta cgtatactcc ggaatattaa
     3901 tagatcatgg agataattaa aatgataacc atctcgcaaa taaataagta ttttactgtt
     3961 ttcgtaacag ttttgtaata aaaaaaccta taaatattcc ggattattca taccgtccca
     4021 ccatcgggcg cggatctcgg tccgaaaacc atgcatcatc atcatcatca cagcagcggc
     4081 agagaaaact tgtatttcca gggccatatg agttctcctc ctgaaagatc cataacttcg
     4141 tatagcatac attatacgaa gttatgcggc cgcgacgtcc acatatacct gccgttcact
     4201 attatttagt gaaatgagat attatgatat tttctgaatt gtgattaaaa aggcaacttt
     4261 atgcccatgc aacagaaact ataaaaaata cagagaatga aaagaaacag atagattttt
     4321 tagttcttta ggcccgtagt ctgcaaatcc ttttatgatt ttctatcaaa caaaagagga
     4381 aaatagacca gttgcaatcc aaacgagagt ctaatagaat gaggtcgaaa agtaaatcgc
     4441 gcgggtttgt tactgataaa gcaggcaaga cctaaaatgt gtaaagggca aagtgtatac
     4501 tttggcgtca ccccttacat attttaggtc tttttttatt gtgcgtaact aacttgccat
     4561 cttcaaacag gagggctgga agaagcagac cgctaacaca gtacataaaa aaggagacat
     4621 gaacgatgaa catcaaaaag tttgcaaaac aagcaacagt attaaccttt actaccgcac
     4681 tgctggcagg aggcgcaact caagcgtttg cgaaagaaac gaaccaaaag ccatataagg
     4741 aaacatacgg catttcccat attacacgcc atgatatgct gcaaatccct gaacagcaaa
     4801 aaaatgaaaa atataaagtt cctgagttcg attcgtccac aattaaaaat atctcttctg
     4861 caaaaggcct ggacgtttgg gacagctggc cattacaaaa cactgacggc actgtcgcaa
     4921 actatcacgg ctaccacatc gtctttgcat tagccggaga tcctaaaaat gcggatgaca
     4981 catcgattta catgttctat caaaaagtcg gcgaaacttc tattgacagc tggaaaaacg
     5041 ctggccgcgt ctttaaagac agcgacaaat tcgatgcaaa tgattctatc ctaaaagacc
     5101 aaacacaaga atggtcaggt tcagccacat ttacatctga cggaaaaatc cgtttattct
     5161 acactgattt ctccggtaaa cattacggca aacaaacact gacaactgca caagttaacg
     5221 tatcagcatc agacagctct ttgaacatca acggtgtaga ggattataaa tcaatctttg
     5281 acggtgacgg aaaaacgtat caaaatgtac agcagttcat cgatgaaggc aactacagct
     5341 caggcgacaa ccatacgctg agagatcctc actacgtaga agataaaggc cacaaatact
     5401 tagtatttga agcaaacact ggaactgaag atggctacca aggcgaagaa tctttattta
     5461 acaaagcata ctatggcaaa agcacatcat tcttccgtca agaaagtcaa aaacttctgc
     5521 aaagcgataa aaaacgcacg gctgagttag caaacggcgc tctcggtatg attgagctaa
     5581 acgatgatta cacactgaaa aaagtgatga aaccgctgat tgcatctaac acagtaacag
     5641 atgaaattga acgcgcgaac gtctttaaaa tgaacggcaa atggtacctg ttcactgact
     5701 cccgcggatc aaaaatgacg attgacggca ttacgtctaa cgatatttac atgcttggtt
     5761 atgtttctaa ttctttaact ggcccataca agccgctgaa caaaactggc cttgtgttaa
     5821 aaatggatct tgatcctaac gatgtaacct ttacttactc acacttcgct gtacctcaag
     5881 cgaaaggaaa caatgtcgtg attacaagct atatgacaaa cagaggattc tacgcagaca
     5941 aacaatcaac gtttgcgcct agcttcctgc tgaacatcaa aggcaagaaa acatctgttg
     6001 tcaaagacag catccttgaa caaggacaat taacagttaa caaataaaaa cgcaaaagaa
     6061 aatgccgata tcctattggc attgacgtca ggtggcactt ttcgaggaga tcatgcacat
     6121 gatgacgaag cttgtcgaga agtactagag gatcataatc agccatacca catttgtaga
     6181 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     6241 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     6301 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     6361 actcatcaat gtatcttatc atgtctggat ctgatcactg atatcgccta ggagatccga
     6421 accagataag tgaaatctag ttccaaacta ttttgtcatt tttaattttc gtattagctt
     6481 acgacgctac acccagttcc catctatttt gtcactcttc cctaaataat ccttaaaaac
     6541 tccatttcca cccctcccag ttcccaacta ttttgtccgc ccacagcggg gcatttttct
     6601 tcctgttatg tttttaatca aacatcctgc caactccatg tgacaaaccg tcatcttcgg
     6661 ctactttttc tctgtcacag aatgaaaatt tttctgtcat ctcttcgtta ttaatgtttg
     6721 taattgactg aatatcaacg cttatttgca gcctgaatgg cgaatgg
//
