LOCUS       sequence-492429-        8050 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Piggybac based mammalian expression vector for histone H2B HaloTag, 
            driven by the EF1a promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8050)
  AUTHORS   Hibino K, Sakai Y, Tamura S, Takagi M, Minami K, Natsume T, Shimazoe
            MA, Kanemaki MT, Imamoto N, Maeshima K
  TITLE     Single-nucleosome imaging unveils that condensins and 
            nucleosome-nucleosome interactions differentially constrain 
            chromatin to organize mitotic chromosomes.
  JOURNAL   Nat Commun. 2024 Aug 21;15(1):7152. doi: 10.1038/s41467-024-51454-y.
  PUBMED    39169041
REFERENCE   2  (bases 1 to 8050)
  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 
            Commun."; date: "2024-08-21"; volume: "15(1):7152. doi"; pages: " 
            10.1038/s41467-024-51454-y"
COMMENT     Sequence Label: pPB-EF1-alpha-H2B-HaloTag-IRES-puro
FEATURES             Location/Qualifiers
     source          1..8050
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             417..685
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     promoter        complement(698..909)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     repeat_region   1362..1424
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(1554..1571)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1554..1570)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1563..1585)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1712..2167
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1799..1818)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2009..2030
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2193..2297
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2298..2366,2367..3158)
                     /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(2516..2535)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3329..3917
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3818..3837
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4071..4088
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4205..4226
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4241..4246,4247..4264,4265..4271)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4279..4295
                     /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     4284..4306
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4303..4319
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4303..4319
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   4810..4844
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     polyA_signal    5607..5728
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5618..5637)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5672..5691
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5770..5790
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5859..6458)
                     /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(5942..5962)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6439..6458
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     misc_feature    join(6471..6479,6480..6482,6483..7057)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(6644..6663)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     6873..6890
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(7095..7979)
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     CDS             complement(join(8046..8050,1..373))
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
ORIGIN
        1 cgctggtgta cttggtgatg gccttagtac cctcggacac ggcgtgcttg gccaactccc
       61 caggcagcag caggcgcacg gccgtctgga tctccctgga ggtgatggtc gagcgcttgt
      121 tgtaatgcgc caggcgggaa gcctcacctg cgatgcgctc gaaaatgtcg ttcacaaacg
      181 aattcatgat gcccatggcc ttggacgaaa tgccggtgtc agggtggacc tgcttcagaa
      241 ccttgtacac atagatggaa tagctctcct tgcggctgcg cttgcgcttc ttgccgcctt
      301 tcttctgcgc cttagtcacc gccttcttgg agcccttttt cggggcggga gcagacttcg
      361 ctggctctgg catctcgaaa ttcgctagct ctagagtagg cgccggtcac agcttggatc
      421 tgtaacggcg cagaacagaa aacgaaacaa agacgtagag ttgagcaagc agggtcaggc
      481 aaagcgtgga gagccggctg agtctaggta ggctccaagg gagcgccgga caaaggcccg
      541 gtctcgacct gagctttaaa cttacctaga cggcggacgc agttcaggag gcaccacagg
      601 cgggaggcgg cagaacgcga ctcaaccggc gtggatggcg gcctcaggta gggcggcggg
      661 cgcgtgaagg agagatgcga gcccctcgaa gcttcagctg tgttctggcg gcaaacccgt
      721 tgcgaaaaag aacgttcacg gcgactactg cacttatata cggttctccc ccaccctcgg
      781 gaaaaaggcg gagccagtac acgacatcac tttcccagtt taccccgcgc caccttctct
      841 aggcacccgt tcaattgccg acccctcccc ccaacttctc ggggactgtg ggcgatgtgc
      901 gctctgccca ctgacgggca ccggagcgat cgcagatcct tggccgtgga ggccttttcc
      961 ccgtatcccc ccaggtgtct gcaggctcaa agagcagcga gaagcgttca gaggaaagcg
     1021 atcccgtgcc accttccccg tgcccgggct gtccccgcac gctgccggct cggggatgcg
     1081 gggggagcgc cggaccggag cggagccccg ggcggctcgc tgctgccccc tagcggggga
     1141 gggacgtaat tacatccctg ggggctttgg gggggggctg tccctcataa aagttttgtt
     1201 actttataga agaaattttg agtttttgtt ttttttaata aataaataaa cataaataaa
     1261 ttgtttgttg aatttattat tagtatgtaa gtgtaaatat aataaaactt aatatctatt
     1321 caaattaata aataaacctc gatatacaga ccgataaaac acatgcgtca attttacaca
     1381 tgattatctt taacgtacgt cacaatatga ttatctttct agggttaatc tagctgcgtg
     1441 ttctgcagcg tgtcgagcat cttcatctgc tccatcacgc tgtaaaacac atttgcaccg
     1501 cgagtctgcc cgtcctccac gggttcaaaa acgtgaatga acgaggcgcg ctcactggcc
     1561 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca
     1621 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc
     1681 caacagttgc gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg
     1741 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     1801 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     1861 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     1921 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     1981 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     2041 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     2101 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt
     2161 acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     2221 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     2281 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     2341 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     2401 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     2461 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     2521 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     2581 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     2641 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     2701 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     2761 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     2821 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     2881 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     2941 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     3001 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     3061 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     3121 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     3181 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     3241 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     3301 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     3361 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     3421 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     3481 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     3541 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     3601 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     3661 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     3721 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     3781 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     3841 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     3901 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     3961 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     4021 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     4081 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
     4141 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
     4201 caattaatgt gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg
     4261 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc
     4321 atgattacgc caagcgcgcc cgccgggtaa ctcacggggt atccatgtcc atttctgcgg
     4381 catccagcca ggatacccgt cctcgctgac gtaatatccc agcgccgcac cgctgtcatt
     4441 aatctgcaca ccggcacggc agttccggct gtcgccggta ttgttcgggt tgctgatgcg
     4501 cttcgggctg accatccgga actgtgtccg gaaaagccgc gacgaactgg tatcccaggt
     4561 ggcctgaacg aacagttcac cgttaaaggc gtgcatggcc acaccttccc gaatcatcat
     4621 ggtaaacgtg cgttttcgct caacgtcaat gcagcagcag tcatcctcgg caaactcttt
     4681 ccatgccgct tcaacctcgc gggaaaaggc acgggcttct tcctccccga tgcccagata
     4741 gcgccagctt gggcgatgac tgagccggaa aaaagacccg acgatatgat cctgatgcag
     4801 ctagattaac cctagaaaga tagtctgcgt aaaattgacg catgcattct tgaaatattg
     4861 ctctctcttt ctaaatagcg cgaatccgtc gctgtgcatt taggacatct cagtcgccgc
     4921 ttggagctcc cgtgaggcgt gcttgtcaat gcggtaagtg tcactgattt tgaactataa
     4981 cgaccgcgtg agtcaaaatg acgcatgatt atcttttacg tgacttttaa gatttaactc
     5041 atacgataat tatattgtta tttcatgttc tacttacgtg ataacttatt atatatatat
     5101 tttcttgtta tagatatcag ggacagcccc cccccaaagc ccccagggat gtaattacgt
     5161 ccctcccccg ctagggggca gcagcgagcc gcccggggct ccgctccggt ccggcgctcc
     5221 ccccgcatcc ccgagccggc agcgtgcggg gacagcccgg gcacggggaa ggtggcacgg
     5281 gatcgctttc ctctgaacgc ttctcgctgc tctttgagcc tgcagacacc tggggggata
     5341 cggggaaaaa gccataccaa tgggccctaa aaaaggaatc cagtcaattc cggggctaaa
     5401 cctggctgcc actgtttctt tagggacttc gttcctgtga ggacaggaaa ccctgcaacc
     5461 tggttgacca gagcaaggga tataagaacc cagtttggga acactgctgt ttaaatatta
     5521 aacagggatg tcttgtcccc cggaagggag accagatgag cttctgatag actaattgac
     5581 atcatttgag tcaattccag acatgataag atacattgat gagtttggac aaaccacaac
     5641 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     5701 aaccattata agctgcaata aacaagttta gttaacgcat gatacaaagg cattaaagca
     5761 gcgtatccac atagcgtaaa aggagcaaca tagttaagaa taccagtcaa tctttcacaa
     5821 attttgtaat ccagaggttg attgtcgaac gcgtcgactc aggcaccggg cttgcgggtc
     5881 atgcaccagg tgcgcggtcc ttcgggcacc tcgacgtcgg cggtgacggt gaagccgagc
     5941 cgctcgtaga aggggaggtt gcggggcgcg gaggtctcca ggaaggcggg caccccggcg
     6001 cgctcggccg cctccactcc ggggagcacg acggcgctgc ccagaccctt gccctggtgg
     6061 tcgggcgaga cgccgacggt ggccaggaac cacgcgggct ccttgggccg gtgcggcgcc
     6121 aggaggcctt ccatctgttg ctgcgcggcc agccgggaac cgctcaactc ggccatgcgc
     6181 gggccgatct cggcgaacac cgcccccgct tcgacgctct ccggcgtggt ccagaccgcc
     6241 accgcggcgc cgtcgtccgc gacccacacc ttgccgatgt cgagcccgac gcgcgtgagg
     6301 aagagttctt gcagctcggt gacccgctcg atgtggcggt ccggatcgac ggtgtggcgc
     6361 gtggcggggt agtcggcgaa cgcggcggcg agggtgcgta cggccctggg gacgtcgtcg
     6421 cgggtggcga ggcgcaccgt gggcttgtac tcggtcattc cggacgccat ggttgtggcc
     6481 atattatcat cgtgtttttc aaaggaaaac cacgtccccg tggttcgggg ggcctagacg
     6541 tttttttaac ctcgactaaa cacatgtaaa gcatgtgcac cgaggcccca gatcagatcc
     6601 catacaatgg ggtaccttct gggcatcctt cagccccttg ttgaatacgc ttgaggagag
     6661 ccatttgact ctttccacaa ctatccaact cacaacgtgg cactggggtt gtgccgcctt
     6721 tgcaggtgta tcttatacac gtggcttttg gccgcagagg cacctgtcgc caggtggggg
     6781 gttccgctgc ctgcaaaggg tcgctacaga cgttgtttgt cttcaagaag cttccagagg
     6841 aactgcttcc ttcacgacat tcaacagacc ttgcattcct ttggcgagag gggaaagacc
     6901 cctaggaatg ctcgtcaaga agacagggcc aggtttccgg gccctcacat tgccaaaaga
     6961 cggcaatatg gtggaaaata acatatagac aaacgcacac cggccttatt ccaagcggct
     7021 tcggccagta acgttagggg ggggggaggg agaggggcgc ggccgcggat ccgatttaaa
     7081 ttcgaattaa cttaaccgga aatctccaga gtagacagcc agcgcgcgat ctcgctgccg
     7141 atcaggtccg ggttgtcttc ttgcagcaga ttcagacccg ggccgatgtc cacagccttg
     7201 cagttaggca ggcttttggc caggcgagcg gcttcggccg gtgggatcag aacgcctggg
     7261 gtgccccaga acagcagctt cgggacaggg gactggtgca gccagtccat gtattcttcg
     7321 accagcgcga cgatgttcgc tggctcaccg gcgattggca gctcgtttgg gaagcgccac
     7381 agtggctcgc ggtcaacagg attcaggaac ggctcgcggt aatggtccat ctcgacttca
     7441 gtcagcgggc ggacgacacc catcggcagc gtaccctcga taaaaacgtt ctgatcgatg
     7501 atcagcttgc ggccgacgtc ggtggtgcgg aaggcctgga aggtctcgcg ggcaaattct
     7561 ggccattcgt cccaggtcgg gatagggcgg atgaactcca taaatgcaat acctttgacg
     7621 cgctctggat tgcgcttggc ccagtggaaa cccagagcgg agccccagtc gtgaatgacc
     7681 aggacgacct cttccagacc cagggcttcg atgaaggcat ccatgaagcg gacgtggtcg
     7741 tcgaagaaat aacccaggtc tggtttgtcg gatttgccca taccgatcag gtctggagca
     7801 atgcagcgat gggtcggtgc aacatgcggg atgatgttgc gccacacgta ggaggaggtc
     7861 gggttaccgt gcaggaacag cacaggggtg ccatcgcgcg gaccaacatc gacgtagtgc
     7921 atgcgctcgc ccaggacttc cacataatgg gggtcgaatg gaaagccagt accgatttcg
     7981 gaaccgccac cagagcctcc acctccactt ccacctccgc ctgaaccacc tccaccagat
     8041 ggatccttag
//
