LOCUS       sequence-292448-        7155 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human RBBP4 in insect cells, , under a C3-cleavable 
            (PreScission Protease) N-terminal hexahistidine-MBP tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7155)
  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 7155)
  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: pFB1.HMBP.PrS.RBBP4
FEATURES             Location/Qualifiers
     source          1..7155
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        296..387
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     312..329
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             448..465
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             472..1569
                     /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      1426..1435
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1543..1566
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1597..1620
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     polyA_signal    3067..3201
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3104..3123)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3158..3177
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  3230..3395
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      3579..4033
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3666..3685)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3875..3896
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4059..4163
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4164..4232,4233..5024)
                     /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(4382..4401)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5195..5783
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5684..5703
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5926..5943
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  6077..6301
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(6368..6901)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(7071..7076,7077..7093,7094..7099))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
ORIGIN
        1 gcagcgaagt cgaggcattt ctgtcctggc tggcgaacga gcgcaaggtt tcggtctcca
       61 cgcatcgtca ggcattggcg gccttgctgt tcttctacgg caaggtgctg tgcacggatc
      121 tgccctggct tcaggagatc ggaagacctc ggccgtcgcg gcgcttgccg gtggtgctga
      181 ccccggatga agtggttcgc atcctcggtt ttctggaagg cgagcatcgt ttgttcgccc
      241 aggactctag ctatagttct agtggttggc tacgtatact ccggaatatt aatagatcat
      301 ggagataatt aaaatgataa ccatctcgca aataaataag tattttactg ttttcgtaac
      361 agttttgtaa taaaaaaacc tataaatatt ccggattatt cataccgtcc caccatcggg
      421 cgcggatctc ggtccgaaac catgggtcat caccatcatc atcacggtac caaaactgaa
      481 gaaggtaaac tggtaatctg gattaacggc gataaaggct ataacggtct cgctgaagtc
      541 ggtaagaaat tcgagaaaga taccggaatt aaagtcaccg ttgagcatcc ggataaactg
      601 gaagagaaat tcccacaggt tgcggcaact ggcgatggcc ctgacattat cttctgggca
      661 cacgaccgct ttggtggcta cgctcaatct ggcctgttgg ctgaaatcac cccggacaaa
      721 gcgttccagg acaagctgta tccgtttacc tgggatgccg tacgttacaa cggcaagctg
      781 attgcttacc cgatcgctgt tgaagcgtta tcgctgattt ataacaaaga tctgctgccg
      841 aacccgccaa aaacctggga agagatcccg gcgctggata aagaactgaa agcgaaaggt
      901 aagagcgcgc tgatgttcaa cctgcaagaa ccgtacttca cctggccgct gattgctgct
      961 gacgggggtt atgcgttcaa gtatgaaaac ggcaagtacg acattaaaga cgtgggcgtg
     1021 gataacgctg gcgcgaaagc gggtctgacc ttcctggttg acctgattaa aaacaaacac
     1081 atgaatgcag acaccgatta ctccatcgca gaagctgcct ttaataaagg cgaaacagcg
     1141 atgaccatca acggcccgtg ggcatggtcc aacatcgaca ccagcaaagt gaattatggt
     1201 gtaacggtac tgccgacctt caagggtcaa ccatccaaac cgttcgttgg cgtgctgagc
     1261 gcaggtatta acgccgccag tccgaacaaa gagctggcaa aagagttcct cgaaaactat
     1321 ctgctgactg atgaaggtct ggaagcggtt aataaagaca aaccgctggg tgccgtagcg
     1381 ctgaagtctt acgaggaaga gttggcgaaa gatccacgta ttgccgccac catggaaaac
     1441 gcccagaaag gtgaaatcat gccgaacatc ccgcagatgt ccgctttctg gtatgccgtg
     1501 cgtactgcgg tgatcaacgc cgccagcggt cgtcagactg tcgatgaagc cctgaaagac
     1561 gcgcagactg gtaccgatta cgatatccca acgaccctgg aagttctgtt ccaggggccc
     1621 ctgggatccg ccgacaagga agcagccttc gacgacgcag tggaagaacg agtgatcaac
     1681 gaggaataca aaatatggaa aaagaacacc ccttttcttt atgatttggt gatgacccat
     1741 gctctggagt ggcccagcct aactgcccag tggcttccag atgtaaccag accagaaggg
     1801 aaagatttca gcattcatcg acttgtcctg gggacacaca catcggatga acaaaaccat
     1861 cttgttatag ccagtgtgca gctccctaat gatgatgctc agtttgatgc gtcacactac
     1921 gacagtgaga aaggagaatt tggaggtttt ggttcagtta gtggaaaaat tgaaatagaa
     1981 atcaagatca accatgaagg agaagtaaac agggcccgtt atatgcccca gaacccttgt
     2041 atcatcgcaa caaagactcc ttccagtgat gttcttgtct ttgactatac aaaacatcct
     2101 tctaaaccag atccttctgg agagtgcaac ccagacttgc gtctccgtgg acatcagaag
     2161 gaaggctatg ggctttcttg gaacccaaat ctcagtgggc acttacttag tgcttcagat
     2221 gaccatacca tctgcctgtg ggacatcagt gccgttccaa aggagggaaa agtggtagat
     2281 gcgaagacca tctttacagg gcatacggca gtagtagaag atgtttcctg gcatctactc
     2341 catgagtctc tgtttgggtc agttgctgat gatcagaaac ttatgatttg ggatactcgt
     2401 tcaaacaata cttccaaacc aagccactca gttgatgctc acactgctga agtgaactgc
     2461 ctttctttca atccttatag tgagttcatt cttgccacag gatcagctga caagactgtt
     2521 gccttgtggg atctgagaaa tctgaaactt aagttgcatt cctttgagtc acataaggat
     2581 gaaatattcc aggttcagtg gtcacctcac aatgagacta ttttagcttc cagtggtact
     2641 gatcgcagac tgaatgtctg ggatttaagt aaaattggag aggaacaatc cccagaagat
     2701 gcagaagacg ggccaccaga gttgttgttt attcatggtg gtcatactgc caagatatct
     2761 gatttctcct ggaatcccaa tgaaccttgg gtgatttgtt ctgtatcaga agacaatatc
     2821 atgcaagtgt ggcaaatggc agagaacatt tataatgatg aagaccctga aggaagcgtg
     2881 gatccagaag gacaagggtc ctgatctaga gcctgcagtc tcgaggcatg cggtaccaag
     2941 cttgtcgaga agtactagag gatcataatc agccatacca catttgtaga ggttttactt
     3001 gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt
     3061 gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     3121 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     3181 gtatcttatc atgtctggat ctgatcactg atatcgccta ggagatccga accagataag
     3241 tgaaatctag ttccaaacta ttttgtcatt tttaattttc gtattagctt acgacgctac
     3301 acccagttcc catctatttt gtcactcttc cctaaataat ccttaaaaac tccatttcca
     3361 cccctcccag ttcccaacta ttttgtccgc ccacagcggg gcatttttct tcctgttatg
     3421 tttttaatca aacatcctgc caactccatg tgacaaaccg tcatcttcgg ctactttttc
     3481 tctgtcacag aatgaaaatt tttctgtcat ctcttcgtta ttaatgtttg taattgactg
     3541 aatatcaacg cttatttgca gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt
     3601 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3661 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3721 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3781 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3841 tcgccctttg acgttggagt ccacgttctt aatagtggac tcttgttcca aactggaaca
     3901 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     3961 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     4021 acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     4081 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     4141 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     4201 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     4261 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     4321 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     4381 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     4441 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     4501 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     4561 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     4621 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     4681 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     4741 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     4801 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     4861 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     4921 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     4981 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     5041 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     5101 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     5161 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     5221 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     5281 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     5341 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     5401 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     5461 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     5521 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     5581 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     5641 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     5701 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     5761 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     5821 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     5881 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     5941 gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     6001 caccgcatag accagccgcg taacctggca aaatcggtta cggttgagta ataaatggat
     6061 gccctgcgta agcgggtgtg ggcggacaat aaagtcttaa actgaacaaa atagatctaa
     6121 actatgacaa taaagtctta aactagacag aatagttgta aactgaaatc agtccagtta
     6181 tgctgtgaaa aagcatactg gacttttgtt atggctaaag caaactcttc attttctgaa
     6241 gtgcaaattg cccgtcgtat taaagagggg cgtggccaag ggcatggtaa agactatatt
     6301 cgcggcgttg tgacaattta ccgaacaact ccgcggccgg gaagccgatc tcggcttgaa
     6361 cgaattgtta ggtggcggta cttgggtcga tatcaaagtg catcacttct tcccgtatgc
     6421 ccaactttgt atagagagcc actgcgggat cgtcaccgta atctgcttgc acgtagatca
     6481 cataagcacc aagcgcgttg gcctcatgct tgaggagatt gatgagcgcg gtggcaatgc
     6541 cctgcctccg gtgctcgccg gagactgcga gatcatagat atagatctca ctacgcggct
     6601 gctcaaactt gggcagaacg taagccgcga gagcgccaac aaccgcttct tggtcgaagg
     6661 cagcaagcgc gatgaatgtc ttactacgga gcaagttccc gaggtaatcg gagtccggct
     6721 gatgttggga gtaggtggct acgtctccga actcacgacc gaaaagatca agagcagccc
     6781 gcatggattt gacttggtca gggccgagcc tacatgtgcg aatgatgccc atacttgagc
     6841 cacctaactt tgttttaggg cgactgccct gctgcgtaac atcgttgctg ctgcgtaaca
     6901 tcaaacatcg acccacggcg taacgcgctt gctgcttgga tgcccgaggc atagactgta
     6961 caaaaaaaca gtcataacaa gccatgaaaa ccgccactgc gccgttacca ccgctgcgtt
     7021 cggtcaaggt tctggaccag ttgcgtgagc gcatacgcta cttgcattac agtttacgaa
     7081 ccgaacaggc ttatgtcaac tgggttcgtg ccttcatccg tttccacggt gtgcgtcacc
     7141 cggcaacctt gggca
//
