LOCUS       sequence-491708-        7018 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  Expresses wild-type H4(Delta-1-19)-Halo under EF1 promoter and can 
            be integrated into FRT site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7018)
  AUTHORS   Semeigazin A, Iida S, Minami K, Tamura S, Ide S, Higashi K, Toyoda 
            A, Kurokawa K, Maeshima K
  TITLE     Behaviors of nucleosomes with mutant histone H4s in euchromatic 
            domains of living human cells.
  JOURNAL   Histochem Cell Biol. 2024 Jul;162(1-2):23-40. doi: 
            10.1007/s00418-024-02293-x. Epub 2024 May 14.
  PUBMED    38743310
REFERENCE   2  (bases 1 to 7018)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 4, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Histochem 
            Cell Biol."; date: "2024-07"; volume: "162(1-2):23-40. doi"; pages: 
            " 10.1007/s00418-024-02293-x. Epub 2024 May 14"
COMMENT     Sequence Label: pEF5-H4-Delta-1-19-HaloTag-FRT
FEATURES             Location/Qualifiers
     source          1..7018
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(735..782)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     914..933
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    1066..1290
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     1279..1296
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(1334..1375)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(1427..2311)
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     primer_bind     2646..2662
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2646..2662
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2701..2720)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2702..2720)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(2737..3915)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2746..3684
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(2769..2789)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     4203..4222
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4297..4401
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4402..4470,4471..5262)
                     /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(4620..4639)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5433..6021
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5922..5941
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6175..6192
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6309..6330
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6345..6350,6351..6368,6369..6375)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6383..6399
                     /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     6388..6410
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6407..6423
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6407..6423
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    6472..6593
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6483..6502)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6537..6556
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(6723..7018,1..727))
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             complement(join(6723..7018,1..727))
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
ORIGIN
        1 gctccataca agccaaccac ggcctccaga agaagatgtt ggcgacctcg tattgggaat
       61 ccccgaacat cgcctcgctc cagtcaatga ccgctgttat gcggccattg tccgtcagga
      121 cattgttgga gccgaaatcc gcgtgcacga ggtgccggac ttcggggcag tcctcggccc
      181 aaagcatcag ctcatcgaga gcctgcgcga cggacgcact gacggtgtcg tccatcacag
      241 tttgccagtg atacacatgg ggatcagcaa tcgcgcatat gaaatcacgc catgtagtgt
      301 attgaccgat tccttgcggt ccgaatgggc cgaacccgct cgtctggcta agatcggccg
      361 cagcgatcgc atccatggcc tccgcgaccg gctgcagaac agcgggcagt tcggtttcag
      421 gcaggtcttg caacgtgaca ccctgtgcac ggcgggagat gcaataggtc aggctctcgc
      481 tgaattcccc aatgtcaagc acttccggaa tcgggagcgc ggccgatgca aagtgccgat
      541 aaacataacg atctttgtag aaaccatcgg cgcagctatt tacccgcagg acatatccac
      601 gccctcctac atcgaagctg aaagcacgag attcttcgcc ctccgagagc tgcatcaggt
      661 cggagacgct gtcgaacttt tcgatcagaa acttctcgac agacgtcgcg gtgagttcag
      721 gctttttggc caaggaagtt cctatacttt ctagagaata ggaacttcgg aataggaact
      781 tctaggtacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
      841 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
      901 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
      961 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     1021 ggggataccc cctagagccc cagctggttc tttccgcctc agaagccata gagcccaccg
     1081 catccccagc atgcctgcta ttgtcttccc aatcctcccc cttgctgtcc tgccccaccc
     1141 caccccccag aatagaatga cacctactca gacaatgcga tgcaatttcc tcattttatt
     1201 aggaaaggac agtgggagtg gcaccttcca gggtcaagga aggcacgggg gaggggcaaa
     1261 caacagatgg ctggcaacta gaaggcacag tcgaggctga tcagcgggtt taaactcatt
     1321 actaaccggt acgcgtagaa tcgagaccga ggagagggtt agggataggc ttaccttcga
     1381 accgcgggcc ctctagactc gagcggccgg ccgccgaatt aacttaaccg gaaatctcca
     1441 gagtagacag ccagcgcgcg atctcgctgc cgatcaggtc cgggttgtct tcttgcagca
     1501 gattcagacc cgggccgatg tccacagcct tgcagttagg caggcttttg gccaggcgag
     1561 cggcttcggc cggtgggatc agaacgcctg gggtgcccca gaacagcagc ttcgggacag
     1621 gggactggtg cagccagtcc atgtattctt cgaccagcgc gacgatgttc gctggctcac
     1681 cggcgattgg cagctcgttt gggaagcgcc acagtggctc gcggtcaaca ggattcagga
     1741 acggctcgcg gtaatggtcc atctcgactt cagtcagcgg gcggacgaca cccatcggca
     1801 gcgtaccctc gataaaaacg ttctgatcga tgatcagctt gcggccgacg tcggtggtgc
     1861 ggaaggcctg gaaggtctcg cgggcaaatt ctggccattc gtcccaggtc gggatagggc
     1921 ggatgaactc cataaatgca atacctttga cgcgctctgg attgcgcttg gcccagtgga
     1981 aacccagagc ggagccccag tcgtgaatga ccaggacgac ctcttccaga cccagggctt
     2041 cgatgaaggc atccatgaag cggacgtggt cgtcgaagaa ataacccagg tctggtttgt
     2101 cggatttgcc cataccgatc aggtctggag caatgcagcg atgggtcggt gcaacatgcg
     2161 ggatgatgtt gcgccacacg taggaggagg tcgggttacc gtgcaggaac agcacagggg
     2221 tgccatcgcg cggaccaaca tcgacgtagt gcatgcgctc gcccaggact tccacataat
     2281 gggggtcgaa tggaaagcca gtaccgattt cggaaccgcc accagagcct ccacctccac
     2341 ttccacctcc gcctgaacca cctccaccag atggatcacc gccaaagcca taaagggtgc
     2401 gtccctggcg cttaagcgcg tagaccacgt ccatggcagt gactgtcttg cgcttggcgt
     2461 gctccgtata ggtgacagcg tcacggatca cgttctccaa aaacaccttg agcaccccgc
     2521 gagtctcctc gtagatcaga ccagagatcc gcttcacacc gccacgccgg gccagacgcc
     2581 ggatggccgg cttggtgatg ccctggatgt tgtcacgcaa caccttcatc tcgaaattcg
     2641 ctagctctag aactagtgga tccgagctcg gtaccaagct tacctagcca gcttgggtct
     2701 ccctatagtg agtcgtatta gtaccaagct aattcctcac gacacctgaa atggaagaaa
     2761 aaaactttga accactgtct gaggcttgag aatgaaccaa gatccaaact caaaaagggc
     2821 aaattccaag gagaattaca tcaagtgcca agctggccta acttcagtct ccacccactc
     2881 agtgtgggga aactccatcg cataaaaccc ctccccccaa cctaaagacg acgtactcca
     2941 aaagctcgag aactaatcga ggtgcctgga cggcgcccgg tactccgtgg agtcacatga
     3001 agcgacggct gaggacggaa aggccctttt cctttgtgtg ggtgactcac ccgcccgctc
     3061 tcccgagcgc cgcgtcctcc attttgagct ccctgcagca gggccgggaa gcggccatct
     3121 ttccgctcac gcaactggtg ccgaccgggc cagccttgcc gcccagggcg gggcgataca
     3181 cggcggcgcg aggccaggca ccagagcagg ccggccagct tgagactacc cccgtccgat
     3241 tctcggtggc cgcgctcgca ggccccgcct cgccgaacat gtgcgctggg acgcacgggc
     3301 cccgtcgccg cccgcggccc caaaaaccga aataccagtg tgcagatctt ggcccgcatt
     3361 tacaagacta tcttgccaga aaaaaagcgt cgcagcaggt catcaaaaat tttaaatggc
     3421 tagagactta tcgaaagcag cgagacaggc gcgaaggtgc caccagattc gcacgcggcg
     3481 gccccagcgc ccaggccagg cctcaactca agcacgaggc gaaggggctc cttaagcgca
     3541 aggcctcgaa ctctcccacc cacttccaac ccgaagctcg ggatcaagaa tcacgtactg
     3601 cagccaggtg gaagtaattc aaggcacgca agggccataa cccgtaaaga ggccaggccc
     3661 gcgggaacca cacacggcac ttacctgtgt tctggcggca aacccgttgc gaaaaagaac
     3721 gttcacggcg actactgcac ttatatacgg ttctccccca ccctcgggaa aaaggcggag
     3781 ccagtacacg acatcacttt cccagtttac cccgcgccac cttctctagg caccggttca
     3841 attgccgacc cctcccccca acttctcggg gactgtgggc gatgtgcgct ctgcccactg
     3901 acgggcaccg gagccaattc ccactccttt caagacctag aaggtccatt agctgcaaag
     3961 attcctctct gtttaaaact ttatccatct ttgcaaagct ttttgcaaaa gcctagtcaa
     4021 taatcaatgt caacgcgtat atctggcccg tacatcgcga agcagcgcaa aacgcctaac
     4081 cctaagcaga ttcttcatgc aattgtcggt caagccttgc cttgttgtag cttaaatttt
     4141 gctcgcgcac tactcagcga cctccaacac acaagcaggg agcagatact ggcttaacta
     4201 tgcggcatca gagcagattg tactgagagt gcaccatagg ggatcgggag atctcccgat
     4261 ccgtcgacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     4321 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     4381 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     4441 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     4501 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     4561 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     4621 ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata
     4681 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     4741 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     4801 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     4861 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     4921 gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact
     4981 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     5041 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     5101 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     5161 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     5221 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
     5281 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     5341 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     5401 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     5461 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     5521 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     5581 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     5641 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     5701 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     5761 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     5821 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     5881 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     5941 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     6001 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     6061 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     6121 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     6181 tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg
     6241 ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc
     6301 aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt
     6361 ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat
     6421 gaccatgatt acgccaagct ctagctagag gtcgacggta tacagacatg ataagataca
     6481 ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa
     6541 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttggggtgg
     6601 gcgaagaact ccagcatgag atccccgcgc tggaggatca tccagccggc gtcccggaaa
     6661 acgattccga agcccaacct ttcatagaag gcggcggtgg aatcgaaatc tcgtagtacg
     6721 tgctattcct ttgccctcgg acgagtgctg gggcgtcggt ttccactatc ggcgagtact
     6781 tctacacagc catcggtcca gacggccgcg cttctgcggg cgatttgtgt acgcccgaca
     6841 gtcccggctc cggatcggac gattgcgtcg catcgaccct gcgcccaagc tgcatcatcg
     6901 aaattgccgt caaccaagct ctgatagagt tggtcaagac caatgcggag catatacgcc
     6961 cggagccgcg gcgatcctgc aagctccgga tgcctccgct cgaagtagcg cgtctgct
//
