LOCUS       sequence-362447-        5692 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Repair template for the C-terminal tagging of H2B histones with 
            mTagBFP in human cells using CRISPR/Cas9..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5692)
  AUTHORS   Husser MC, Ozugergin I, Resta T, Martin VJJ, Piekny AJ
  TITLE     Cytokinetic diversity in mammalian cells is revealed by the 
            characterization of endogenous anillin, Ect2 and RhoA.
  JOURNAL   Open Biol. 2022 Nov;12(11):220247. doi: 10.1098/rsob.220247. Epub 
            2022 Nov 23.
  PUBMED    36416720
REFERENCE   2  (bases 1 to 5692)
  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: "Open 
            Biol."; date: "2022-11"; volume: "12(11):220247. doi"; pages: " 
            10.1098/rsob.220247. Epub 2022 Nov 23"
COMMENT     Sequence Label: pODN-H2BC11-TagBFP
FEATURES             Location/Qualifiers
     source          1..5692
                     /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     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    880..901
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(916..921,922..939,940..946)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2003..2380
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKDGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAVTKYTSAK"
     primer_bind     complement(4146..4165)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4147..4165)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4471..4489)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4557..4661
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4662..4730,4731..5522)
                     /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(4880..4899)
                     /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 ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcaattg gcagtgagcg caacgcaatt aatgtgagtt agctcactca
      901 ttaggcaccc caggctttac actttatgct tccggctcgt ataatgtgtg gaattatgag
      961 cggataataa tttcacacag gaggtttaaa ctttaaacat gtcaaaagag acgtcttttg
     1021 ttaagaatgc tgaggaactt gcaaagcaaa aaatggatgc tattaaccct gaactttctt
     1081 caaaatttaa atttttaata aaattcctgt ctcagtttcc tgaagcttgc tctaaacctc
     1141 gttcaaaaaa aatgcagaat aaagttggtc aagaggaaca tattgaatat ttagctcgta
     1201 gttttcatga gagtcgattg ccaagaaaac ccacgccacc tacaacggtt cctgatgagg
     1261 tggttagcat agttcttaat ataagtttta atatacagcc tgaaaatctt gagagaataa
     1321 aagaagaaca tcgattttcc atggcagctg agaatattgt aggagatctt ctagaaagat
     1381 ctttgcccag cagcttgttg agctcctcgt cgttgcggat ggccagttgg agatgacgcg
     1441 ggatgatgcg ggtcttcttg ttgtcgcggg ccgcgttgcc cgccagttcc aggatctcgg
     1501 cggtcaggta ctccagcacc gctgccagat acaccggcgc gccggccccg acccgctcgg
     1561 catagttgcc tttgcggagc aggcggtgca ctcggcccac ggggaactgg agaccggccc
     1621 tagaagagcg agtcttggcc ttagcgcggg ctttgcctcc ctgcttgcca cgtccagaca
     1681 tagcgagcgc aactcactac gagcaaccac aaagtgaacg ggaaaggcgg cgctttttat
     1741 aaacactatt gggcgcgaaa aagaagacgt gttgttggtt agggctgcag tttaatttca
     1801 accaatagta gtgcgtcttc tggatttgcg aatcctgatt gggcagacct gacctctgac
     1861 gttaccctga ataactacca atcagacaca agacttcaac tcttcacctt atttgcataa
     1921 gcgattctat ataaaagcgc cttgtcatac cctgctcacg ctgtttttcc ttttcgttgg
     1981 cgctttatag ctacacagtg ctatgccaga gccagcgaag tctgctcccg ccccgaaaaa
     2041 gggctccaag aaggcggtga ctaaggcgca gaagaaagac ggcaagaagc gcaagcgcag
     2101 ccgcaaggag agctattcca tctatgtgta caaggttctg aagcaggtcc accctgacac
     2161 cggcatttcg tccaaggcca tgggcatcat gaattcgttt gtgaacgaca ttttcgagcg
     2221 catcgcaggt gaggcttccc gcctggcgca ttacaacaag cgctcgacca tcacctccag
     2281 ggagatccag acggccgtgc gcctgctgct gcctggggag ttggccaagc acgccgtgtc
     2341 cgagggtact aaggccgtca ccaagtacac cagcgctaag gaccctcctg tggccaccat
     2401 gagcgaactg atcaaagaga acatgcacat gaagctgtac atggaaggca ccgttgacaa
     2461 ccaccacttt aagtgcacgt ctgagggtga gggtaagccg tacgaaggca cccaaaccat
     2521 gcgtatcaaa gttgtggagg gcggtccact gccgttcgct tttgacattc tggcgaccag
     2581 cttcctgtac ggttccaaaa cgttcattaa ccatactcag ggcattccgg atttcttcaa
     2641 acagagcttt ccggaaggtt tcacctggga gcgtgtcacc acgtatgaag atggtggtgt
     2701 gttgaccgcc acccaagata cctccctgca agatggctgt ctgatctata acgtgaaaat
     2761 tcgtggcgtc aactttacga gcaatggtcc ggtgatgcag aagaaaaccc tgggttggga
     2821 ggcgtttacg gaaaccctgt atccggccga tggtggcctg gagggccgta acgacatggc
     2881 actgaagctg gttggtggca gccatttgat cgcaaatatc aagacgacgt accgcagcaa
     2941 gaaaccggcg aaaaatctga agatgccggg tgtttactat gtcgactacc gtctggaacg
     3001 cattaaagaa gcgaataatg agacttacgt ggagcagcac gaggttgcag tcgcgcgcta
     3061 ttgcgacttg cctagcaagc tgggtcataa actgaattaa acagtgagtt ggttgcaaac
     3121 tctcaatcct aacgcctctt ttaagagcca cccatgttct caaagaaaga gctggtgctt
     3181 gtattcctcc tctgctggcc actgacaaac ccttgtaact tgctactgtg ttttttggtc
     3241 tgaagtagag cagttattta actaatcctt agtgactttt tttttttaga tctgccattc
     3301 taatcttaga gttaagtaag gagatgggaa attttctatt ataagttcga aaccaattaa
     3361 aatacgttag aaaccaatta aaatactcgt cggtcccccg tcggttagtg atttggaaca
     3421 gtgccaagtt gcagcggttg tcagtttgaa tttgcccggg caacgcccgc ccttcctctg
     3481 catcctgatt ggttgttgta tccaggaagt ccacggtcgg cttgggtgcg ccaaaggaat
     3541 ccaaaccccg cctttatttg gccttcctta caaactcagt aaactgtcag tttctcattc
     3601 tttagtattt tatcgctttc gggaaggaga tttatttcta attagttttc gttggataga
     3661 gcgttttttt ttttaaaggg accctttatg gcaacataat cagttttttt tttttttgag
     3721 aaggagactt cctctgtcgc ccagactgga gtgcggtggc gcgatctcag ctcactgcag
     3781 cctccgcctc ccgggttcca gcgattctcc tgcttcaccc ttccgtgtag ctgggattac
     3841 aggcatgcgc caccatgccc ggataatttt tgtattttta gtagagatgg ggtttcacca
     3901 tgttggctag gctggtctcg aactgacttc aggtgatcca ctagcctcag ccttaaagtg
     3961 ctaggattac aggcatgagc catcaagccc ggctacaacg gaatttgaat gctgtagttt
     4021 tcagcctgag aaagatcaat gctttatttc tgattagaat actattttcc ctttatggta
     4081 tatctatata agcaaaggat cttgctgaaa aactcgagcc atccggaaga tctggcggcc
     4141 gctctcccta tagtgagtcg tattacgccg gatggatatg gtgttcaggc acaagtgtta
     4201 aagcagttga ttttattcac tatgatgaaa aaaacaatga atggaacctg ctccaagtta
     4261 aaaatagaga taataccgaa aactcatcga gtagtaagat tagagataat acaacaataa
     4321 aaaaatggtt tagaacttac tcacagcgtg atgctactaa ttgggacaat tttccagatg
     4381 aagtatcatc taagaattta aatgaagaag acttcagagc ttttgttaaa aattatttgg
     4441 caaaaataat ataattcggc tgcaggggcg gcctcgtgat acgcctattt ttataggtta
     4501 atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg
     4561 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     4621 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     4681 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     4741 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     4801 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     4861 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     4921 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     4981 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     5041 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     5101 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     5161 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
     5221 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     5281 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     5341 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     5401 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     5461 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     5521 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     5581 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     5641 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tc
//
