LOCUS       sequence-449676-        7036 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a human CBX8 mutant (KR21A) 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 7036)
  AUTHORS   Uckelmann M, Levina V, Taveneau C, Han Ng X, Pandey V, Martinez J, 
            Mendiratta S, Houx J, Boudes M, Venugopal H, Trepout S, Zhang Q, 
            Flanigan S, Li M, Sierecki E, Gambin Y, Das PP, Bell O, de Marco A, 
            Davidovich C
  TITLE     Dynamic PRC1-CBX8 stabilizes a porous structure of chromatin 
            condensates.
  JOURNAL   bioRxiv [Preprint]. 2024 Feb 12:2023.05.08.539931. doi: 
            10.1101/2023.05.08.539931.
  PUBMED    38405976
REFERENCE   2  (bases 1 to 7036)
  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: "bioRxiv 
            [Preprint]. 2024 Feb 12:2023.05.08.539931. doi: 
            10.1101/2023.05.08.539931."
COMMENT     Sequence Label: pFB1.HMBP.PrS.CBX8-KR21A
FEATURES             Location/Qualifiers
     source          1..7036
                     /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        2255..2346
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     2271..2288
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             2407..2424
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2431..3528
                     /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      3385..3394
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     3502..3525
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             3565..3588
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     polyA_signal    4909..5043
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4946..4965)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5000..5019
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  5072..5237
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      5421..5875
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5508..5527)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5717..5738
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5901..6005
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6006..6074,6075..6866)
                     /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(6224..6243)
                     /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 ctacatactc cggaatatta atagatcatg gagataatta aaatgataac
     2281 catctcgcaa ataaataagt attttactgt tttcgtaaca gttttgtaat aaaaaaacct
     2341 ataaatattc cggattattc ataccgtccc accatcgggc gcggatctcg gtccgaaacc
     2401 atgggtcatc accatcatca tcacggtacc aaaactgaag aaggtaaact ggtaatctgg
     2461 attaacggcg ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat
     2521 accggaatta aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt
     2581 gcggcaactg gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac
     2641 gctcaatctg gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat
     2701 ccgtttacct gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt
     2761 gaagcgttat cgctgattta taacaaagat ctgctgccga acccgccaaa aacctgggaa
     2821 gagatcccgg cgctggataa agaactgaaa gcgaaaggta agagcgcgct gatgttcaac
     2881 ctgcaagaac cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag
     2941 tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg
     3001 ggtctgacct tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac
     3061 tccatcgcag aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg
     3121 gcatggtcca acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc
     3181 aagggtcaac catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt
     3241 ccgaacaaag agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg
     3301 gaagcggtta ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag
     3361 ttggcgaaag atccacgtat tgccgccacc atggaaaacg cccagaaagg tgaaatcatg
     3421 ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc
     3481 gccagcggtc gtcagactgt cgatgaagcc ctgaaagacg cgcagactgc ggccgcaggc
     3541 accgattacg atatcccaac gaccctggaa gttctgttcc aggggcccgg gatggaatta
     3601 tcagcagtag gtgaacgcgt gtttgcagcg gaagcgctgc tcaaacgtag gattcggaag
     3661 ggtcgcatgg agtacctagt taagtggaaa ggatggtcac agaagtatag cacatgggaa
     3721 ccggaggaga acatactgga tgctcgcttg ctagcagcgt ttgaggagcg ggaacgagaa
     3781 atggagctgt acggacctaa aaaacgtgga cccaagccaa agaccttcct tttgaaggct
     3841 caagcggctg ccaaggctgc cacgtatgaa ttcagatcgg attctgccgc aggtataaga
     3901 attccctacc caggggccag tccacaggac ttggcctcca cgtctagggc agcagagggc
     3961 ttacggaaca tgggcctctc tcctccggct tcatctacta gtacgagtag tacatgtgcc
     4021 gcggaagcgc caagggacgc agaccgagac gcagatcggg atgccgagag agatgcggaa
     4081 cgagaggcgg aaagagaagc cgagcgcgag gctgagcgtg aagcggaacg tggcacaagc
     4141 gccgttgatg acaaaccatc gagccctggt gattccagca aaaaacgggg acccaagcct
     4201 aggaaggagt tgccggaccc atcccaagcc ccgcttggtg aaccatcggc gggcctcggg
     4261 gaatatctta aaggccgagc tttggacgat acccctagtg gtgcaggaaa atttcctgcg
     4321 ggacattcgg ttatccaact tgctgcacga caagactcag atttagtaca gtgcggggtg
     4381 acatccccca gctctgcaga ggcgaccggg gccctagctg tcgacacctt cccagcacgc
     4441 gtgatagcgc accgagccgc atttttagaa gctgctgggc agggcgcgct agatcccaac
     4501 ggcactaggg tgcggcacgg ttcaggacct ccctcgtccg gggggggcct atatagagac
     4561 atgggagctc aggggggtag accgtcgctt atcgctcgta tcccggtagc ccgtattctg
     4621 ggtgacccgg aggaagaatc ctggtctccc agtttaacga atttagagaa agtcgtagtc
     4681 actgacgtta cgtcaaattt tctgacagtc accatcaaag agagcaatac tgatcaagga
     4741 ttcttcaagg aaaagcgcta actcgaggca tgcggtacca agcttgtcga gaagtactag
     4801 aggatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
     4861 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
     4921 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     4981 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg
     5041 atctgatcac tgatatcgcc taggagatcc gaaccagata agtgaaatct agttccaaac
     5101 tattttgtca tttttaattt tcgtattagc ttacgacgct acacccagtt cccatctatt
     5161 ttgtcactct tccctaaata atccttaaaa actccatttc cacccctccc agttcccaac
     5221 tattttgtcc gcccacagcg gggcattttt cttcctgtta tgtttttaat caaacatcct
     5281 gccaactcca tgtgacaaac cgtcatcttc ggctactttt tctctgtcac agaatgaaaa
     5341 tttttctgtc atctcttcgt tattaatgtt tgtaattgac tgaatatcaa cgcttatttg
     5401 cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     5461 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     5521 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     5581 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     5641 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     5701 gtccacgttc ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     5761 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     5821 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg
     5881 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     5941 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     6001 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     6061 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     6121 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     6181 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     6241 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     6301 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     6361 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     6421 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     6481 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     6541 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     6601 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     6661 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     6721 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     6781 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     6841 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     6901 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     6961 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     7021 agatcaaagg atcttc
//
