LOCUS       sequence-491707-        7120 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  Expresses wild-type H4(Q5,8,12,16)-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 7120)
  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 7120)
  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(Q5,8,12,16)-HaloTag-FRT
FEATURES             Location/Qualifiers
     source          1..7120
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     517..536
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        611..715
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(716..784,785..1576)
                     /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(934..953)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1747..2335
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2236..2255
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2489..2506
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2623..2644
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2659..2664,2665..2682,2683..2689)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2697..2713
                     /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     2702..2724
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2721..2737
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2721..2737
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    2786..2907
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2797..2816)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2851..2870
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(3037..4059)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             complement(3037..4059)
                     /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
                     "
     protein_bind    complement(4067..4114)
                     /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     4246..4265
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    4398..4622
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     4611..4628
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(4666..4707)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(4759..5643)
                     /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     complement(6135..6154)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6136..6154)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(join(6171..7120,1..229))
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          6180..7118
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(6203..6223)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
ORIGIN
        1 gtgttctggc ggcaaacccg ttgcgaaaaa gaacgttcac ggcgactact gcacttatat
       61 acggttctcc cccaccctcg ggaaaaaggc ggagccagta cacgacatca ctttcccagt
      121 ttaccccgcg ccaccttctc taggcaccgg ttcaattgcc gacccctccc cccaacttct
      181 cggggactgt gggcgatgtg cgctctgccc actgacgggc accggagcca attcccactc
      241 ctttcaagac ctagaaggtc cattagctgc aaagattcct ctctgtttaa aactttatcc
      301 atctttgcaa agctttttgc aaaagcctag tcaataatca atgtcaacgc gtatatctgg
      361 cccgtacatc gcgaagcagc gcaaaacgcc taaccctaag cagattcttc atgcaattgt
      421 cggtcaagcc ttgccttgtt gtagcttaaa ttttgctcgc gcactactca gcgacctcca
      481 acacacaagc agggagcaga tactggctta actatgcggc atcagagcag attgtactga
      541 gagtgcacca taggggatcg ggagatctcc cgatccgtcg acgtcaggtg gcacttttcg
      601 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
      661 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
      721 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
      781 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
      841 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
      901 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
      961 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     1021 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     1081 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     1141 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     1201 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     1261 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     1321 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     1381 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     1441 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     1501 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     1561 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     1621 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     1681 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     1741 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     1801 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     1861 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     1921 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     1981 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     2041 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     2101 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     2161 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     2221 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     2281 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     2341 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     2401 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     2461 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     2521 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     2581 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac
     2641 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
     2701 gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca agctctagct
     2761 agaggtcgac ggtatacaga catgataaga tacattgatg agtttggaca aaccacaact
     2821 agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta
     2881 accattataa gctgcaataa acaagttggg gtgggcgaag aactccagca tgagatcccc
     2941 gcgctggagg atcatccagc cggcgtcccg gaaaacgatt ccgaagccca acctttcata
     3001 gaaggcggcg gtggaatcga aatctcgtag tacgtgctat tcctttgccc tcggacgagt
     3061 gctggggcgt cggtttccac tatcggcgag tacttctaca cagccatcgg tccagacggc
     3121 cgcgcttctg cgggcgattt gtgtacgccc gacagtcccg gctccggatc ggacgattgc
     3181 gtcgcatcga ccctgcgccc aagctgcatc atcgaaattg ccgtcaacca agctctgata
     3241 gagttggtca agaccaatgc ggagcatata cgcccggagc cgcggcgatc ctgcaagctc
     3301 cggatgcctc cgctcgaagt agcgcgtctg ctgctccata caagccaacc acggcctcca
     3361 gaagaagatg ttggcgacct cgtattggga atccccgaac atcgcctcgc tccagtcaat
     3421 gaccgctgtt atgcggccat tgtccgtcag gacattgttg gagccgaaat ccgcgtgcac
     3481 gaggtgccgg acttcggggc agtcctcggc ccaaagcatc agctcatcga gagcctgcgc
     3541 gacggacgca ctgacggtgt cgtccatcac agtttgccag tgatacacat ggggatcagc
     3601 aatcgcgcat atgaaatcac gccatgtagt gtattgaccg attccttgcg gtccgaatgg
     3661 gccgaacccg ctcgtctggc taagatcggc cgcagcgatc gcatccatgg cctccgcgac
     3721 cggctgcaga acagcgggca gttcggtttc aggcaggtct tgcaacgtga caccctgtgc
     3781 acggcgggag atgcaatagg tcaggctctc gctgaattcc ccaatgtcaa gcacttccgg
     3841 aatcgggagc gcggccgatg caaagtgccg ataaacataa cgatctttgt agaaaccatc
     3901 ggcgcagcta tttacccgca ggacatatcc acgccctcct acatcgaagc tgaaagcacg
     3961 agattcttcg ccctccgaga gctgcatcag gtcggagacg ctgtcgaact tttcgatcag
     4021 aaacttctcg acagacgtcg cggtgagttc aggctttttg gccaaggaag ttcctatact
     4081 ttctagagaa taggaacttc ggaataggaa cttctaggta cgtgaaccat caccctaatc
     4141 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     4201 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     4261 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     4321 cgccgcgctt aatgcgccgc tacagggcgc gtggggatac cccctagagc cccagctggt
     4381 tctttccgcc tcagaagcca tagagcccac cgcatcccca gcatgcctgc tattgtcttc
     4441 ccaatcctcc cccttgctgt cctgccccac cccacccccc agaatagaat gacacctact
     4501 cagacaatgc gatgcaattt cctcatttta ttaggaaagg acagtgggag tggcaccttc
     4561 cagggtcaag gaaggcacgg gggaggggca aacaacagat ggctggcaac tagaaggcac
     4621 agtcgaggct gatcagcggg tttaaactca ttactaaccg gtacgcgtag aatcgagacc
     4681 gaggagaggg ttagggatag gcttaccttc gaaccgcggg ccctctagac tcgagcggcc
     4741 ggccgccgaa ttaacttaac cggaaatctc cagagtagac agccagcgcg cgatctcgct
     4801 gccgatcagg tccgggttgt cttcttgcag cagattcaga cccgggccga tgtccacagc
     4861 cttgcagtta ggcaggcttt tggccaggcg agcggcttcg gccggtggga tcagaacgcc
     4921 tggggtgccc cagaacagca gcttcgggac aggggactgg tgcagccagt ccatgtattc
     4981 ttcgaccagc gcgacgatgt tcgctggctc accggcgatt ggcagctcgt ttgggaagcg
     5041 ccacagtggc tcgcggtcaa caggattcag gaacggctcg cggtaatggt ccatctcgac
     5101 ttcagtcagc gggcggacga cacccatcgg cagcgtaccc tcgataaaaa cgttctgatc
     5161 gatgatcagc ttgcggccga cgtcggtggt gcggaaggcc tggaaggtct cgcgggcaaa
     5221 ttctggccat tcgtcccagg tcgggatagg gcggatgaac tccataaatg caataccttt
     5281 gacgcgctct ggattgcgct tggcccagtg gaaacccaga gcggagcccc agtcgtgaat
     5341 gaccaggacg acctcttcca gacccagggc ttcgatgaag gcatccatga agcggacgtg
     5401 gtcgtcgaag aaataaccca ggtctggttt gtcggatttg cccataccga tcaggtctgg
     5461 agcaatgcag cgatgggtcg gtgcaacatg cgggatgatg ttgcgccaca cgtaggagga
     5521 ggtcgggtta ccgtgcagga acagcacagg ggtgccatcg cgcggaccaa catcgacgta
     5581 gtgcatgcgc tcgcccagga cttccacata atgggggtcg aatggaaagc cagtaccgat
     5641 ttcggaaccg ccaccagagc ctccacctcc acttccacct ccgcctgaac cacctccacc
     5701 agatggatca ccgccaaagc cataaagggt gcgtccctgg cgcttaagcg cgtagaccac
     5761 gtccatggca gtgactgtct tgcgcttggc gtgctccgta taggtgacag cgtcacggat
     5821 cacgttctcc aaaaacacct tgagcacccc gcgagtctcc tcgtagatca gaccagagat
     5881 ccgcttcaca ccgccacgcc gggccagacg ccggatggcc ggcttggtga tgccctggat
     5941 gttgtcacgc aacaccttgc ggtggcgctg ggcacccccc tgacccaaac cctgcccgcc
     6001 ctgaccacgt ccagacatct cgaaattcgc tagctctaga gcggccgcca ctgtgctgga
     6061 tatctgcaga attccaccac actggactag tggatccgag ctcggtacca agcttaccta
     6121 gccagcttgg gtctccctat agtgagtcgt attagtacca agctaattcc tcacgacacc
     6181 tgaaatggaa gaaaaaaact ttgaaccact gtctgaggct tgagaatgaa ccaagatcca
     6241 aactcaaaaa gggcaaattc caaggagaat tacatcaagt gccaagctgg cctaacttca
     6301 gtctccaccc actcagtgtg gggaaactcc atcgcataaa acccctcccc ccaacctaaa
     6361 gacgacgtac tccaaaagct cgagaactaa tcgaggtgcc tggacggcgc ccggtactcc
     6421 gtggagtcac atgaagcgac ggctgaggac ggaaaggccc ttttcctttg tgtgggtgac
     6481 tcacccgccc gctctcccga gcgccgcgtc ctccattttg agctccctgc agcagggccg
     6541 ggaagcggcc atctttccgc tcacgcaact ggtgccgacc gggccagcct tgccgcccag
     6601 ggcggggcga tacacggcgg cgcgaggcca ggcaccagag caggccggcc agcttgagac
     6661 tacccccgtc cgattctcgg tggccgcgct cgcaggcccc gcctcgccga acatgtgcgc
     6721 tgggacgcac gggccccgtc gccgcccgcg gccccaaaaa ccgaaatacc agtgtgcaga
     6781 tcttggcccg catttacaag actatcttgc cagaaaaaaa gcgtcgcagc aggtcatcaa
     6841 aaattttaaa tggctagaga cttatcgaaa gcagcgagac aggcgcgaag gtgccaccag
     6901 attcgcacgc ggcggcccca gcgcccaggc caggcctcaa ctcaagcacg aggcgaaggg
     6961 gctccttaag cgcaaggcct cgaactctcc cacccacttc caacccgaag ctcgggatca
     7021 agaatcacgt actgcagcca ggtggaagta attcaaggca cgcaagggcc ataacccgta
     7081 aagaggccag gcccgcggga accacacacg gcacttacct
//
