LOCUS       sequence-447909-        5843 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mutation of valine 192 to phenylalanine (V192F) in macroH2A2..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5843)
  AUTHORS   Arroyo M, Casas-Delucchi CS, Pabba MK, Prorok P, Pradhan SK, Rausch 
            C, Lehmkuhl A, Maiser A, Buschbeck M, Pasque V, Bernstein E, Luck K,
            Cardoso MC
  TITLE     Histone variant macroH2A1 regulates synchronous firing of 
            replication origins in the inactive X chromosome.
  JOURNAL   Nucleic Acids Res. 2024 Aug 27:gkae734. doi: 10.1093/nar/gkae734.
  PUBMED    39189450
REFERENCE   2  (bases 1 to 5843)
  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: "Nucleic 
            Acids Res. 2024 Aug 27:gkae734. doi: 10.1093/nar/gkae734."
COMMENT     Sequence Label: macroH2A2-V192F-GFP (pc5116)
FEATURES             Location/Qualifiers
     source          1..5843
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(1..589)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(81..100)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    918..965
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     990..1009
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(1197..1991)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(1197..1991)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1308..1327)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1918..1937
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(2026..2383)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2040..2175
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        2192..2383
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     2340..2360
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(2385..2489)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2516..2971
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2603..2622)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2813..2834
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    2978..3099
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2989..3008)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3043..3062
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(3222..3935,3936..3938,3939..3941))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(3225..3935,3936..3938,3939..3941))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3267..3288)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     3616..3635
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3875..3896
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3938..3947
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(5162..5365)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(5191..5211)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5366..5669
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
       61 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
      121 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
      181 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
      241 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
      301 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
      361 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
      421 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
      481 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
      541 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
      601 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
      661 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
      721 atcaatctaa agtatatatg agtaacctga ggctatggca gggcctgccg ccccgacgtt
      781 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac
      841 gcgggcgtat tggccccaat ggggtctcgg tggggtatcg acagagtgcc agccctggga
      901 ccgaaccccg cgtttatgaa caaacgaccc aacaccgtgc gttttattct gtctttttat
      961 tgccgtcata gcgcgggttc cttccggtat tgtctccttc cgtgtttcag ttagcctccc
     1021 cctagggtgg gcgaagaact ccagcatgag atccccgcgc tggaggatca tccagccggc
     1081 gtcccggaaa acgattccga agcccaacct ttcatagaag gcggcggtgg aatcgaaatc
     1141 tcgtgatggc aggttgggcg tcgcttggtc ggtcatttcg aaccccagag tcccgctcag
     1201 aagaactcgt caagaaggcg atagaaggcg atgcgctgcg aatcgggagc ggcgataccg
     1261 taaagcacga ggaagcggtc agcccattcg ccgccaagct cttcagcaat atcacgggta
     1321 gccaacgcta tgtcctgata gcggtccgcc acacccagcc ggccacagtc gatgaatcca
     1381 gaaaagcggc cattttccac catgatattc ggcaagcagg catcgccatg ggtcacgacg
     1441 agatcctcgc cgtcgggcat gctcgccttg agcctggcga acagttcggc tggcgcgagc
     1501 ccctgatgct cttcgtccag atcatcctga tcgacaagac cggcttccat ccgagtacgt
     1561 gctcgctcga tgcgatgttt cgcttggtgg tcgaatgggc aggtagccgg atcaagcgta
     1621 tgcagccgcc gcattgcatc agccatgatg gatactttct cggcaggagc aaggtgagat
     1681 gacaggagat cctgccccgg cacttcgccc aatagcagcc agtcccttcc cgcttcagtg
     1741 acaacgtcga gcacagctgc gcaaggaacg cccgtcgtgg ccagccacga tagccgcgct
     1801 gcctcgtctt gcagttcatt cagggcaccg gacaggtcgg tcttgacaaa aagaaccggg
     1861 cgcccctgcg ctgacagccg gaacacggcg gcatcagagc agccgattgt ctgttgtgcc
     1921 cagtcatagc cgaatagcct ctccacccaa gcggccggag aacctgcgtg caatccatct
     1981 tgttcaatca tgcgaaacga tcctcatcct gtctcttgat cgatctttgc aaaagcctag
     2041 gcctccaaaa aagcctcctc actacttctg gaatagctca gaggccgagg cggcctcggc
     2101 ctctgcataa ataaaaaaaa ttagtcagcc atggggcgga gaatgggcgg aactgggcgg
     2161 agttaggggc gggatgggcg gagttagggg cgggactatg gttgctgact aattgagatg
     2221 catgctttgc atacttctgc ctgctgggga gcctggggac tttccacacc tggttgctga
     2281 ctaattgaga tgcatgcttt gcatacttct gcctgctggg gagcctgggg actttccaca
     2341 ccctaactga cacacattcc acagctggtt ctttccgcct caggactctt cctttttcaa
     2401 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     2461 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgcg
     2521 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     2581 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     2641 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     2701 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     2761 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     2821 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     2881 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     2941 aattttaaca aaatattaac gcttacaatt tacgccttaa gatacattga tgagtttgga
     3001 caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt
     3061 gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa ttgcattcat
     3121 tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta aaacctctac
     3181 aaatgtggta tggctgatta tgatctagag tcgcggccgc tttacttgta cagctcgtcc
     3241 atgccgagag tgatcccggc ggcggtcacg aactccagca ggaccatgtg atcgcgcttc
     3301 tcgttggggt ctttgctcag ggcggactgg gtgctcaggt agtggttgtc gggcagcagc
     3361 acggggccgt cgccgatggg ggtgttctgc tggtagtggt cggcgagctg cacgctgccg
     3421 tcctcgatgt tgtggcggat cttgaagttc accttgatgc cgttcttctg cttgtcggcc
     3481 atgatataga cgttgtggct gttgtagttg tactccagct tgtgccccag gatgttgccg
     3541 tcctccttga agtcgatgcc cttcagctcg atgcggttca ccagggtgtc gccctcgaac
     3601 ttcacctcgg cgcgggtctt gtagttgccg tcgtccttga agaagatggt gcgctcctgg
     3661 acgtagcctt cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg gtcggggtag
     3721 cggctgaagc actgcacgcc gtaggtcagg gtggtcacga gggtgggcca gggcacgggc
     3781 agcttgccgg tggtgcagat gaacttcagg gtcagcttgc cgtaggtggc atcgccctcg
     3841 ccctcgccgg acacgctgaa cttgtggccg tttacgtcgc cgtccagctc gaccaggatg
     3901 ggcaccaccc cggtgaacag ctcctcgccc ttgctcacca tggtggcgac cggtggatcc
     3961 cgggcccgcg gtaccgtcga ctgcagcttg gcgtcgagct tggccatctc ctgcacgtag
     4021 atgccgatgc tctcgctgtc gaagagcagg aagtacacgt tcttcagcga ggacgcgctc
     4081 gagtcatcaa agtgggctga gatggctttg agggtcacct gggccgcagt ctgtttggga
     4141 aagcagtttc tgccgctggg gaaaggcggg aacgcgacgg actttagctt cttgtcctcc
     4201 gccgctgaca ggcagttttt gatggtctct tcaagctgtt cttcacattt gtcggagccc
     4261 cactgaggga tgtgacagtg gatgacaaat ttggctgcga gtccactgga ttggctgacg
     4321 gcggcttcgg cgacttccaa agggccttgg gatttgcgaa gctcctttac cgtttccaag
     4381 aactcttttc ccccagcctt ttccaaggct ttacctatat cttctttgag gtcaatttcg
     4441 gctgtggttg ggtggacaat gccctccact ctcatggagc caatatggct gatgtcactc
     4501 tgggttaagg acagcttctg tcccagaaaa aggctcttag aagacagaat ggtgaatcca
     4561 tcccctggcc catcttcaga ggtggaattt gaagttcctt ctttatcgct gtcctttggt
     4621 ttggactttt tggacgtccg tggtttggca gccttggatt tcttcccccc cttcttgcct
     4681 gacgtggcct tcctgcctct tttctctggg ggtggggaga ggatcgtttc cgacttgcct
     4741 ttggtccctc gctttttggc cagcagttcg gggtgaattc tgggcaggac gcctccactg
     4801 gcgatggtca ctccttttag cagctggttg agctcctcgt cattggcaac tgccagcaag
     4861 atgtgtctcg gggctatccg ggccttcttg ttgtccctcg cggcattgcc ggccaattct
     4921 agaatttccg ctgccaggta ctcaatgact gccgccatgt agacaggggc gcccacgctg
     4981 atccggtact tgaacgtccc tttcttcaga taacgcatca gcctccccac tggaaagatg
     5041 acacctgccc tagctgaacg ggacagcttg gacattttct tcttcccact ccggcccgac
     5101 ataagcttga gctcgagatc tgagtccggt agcgctagcg gatctgacgg ttcactaaac
     5161 cagctctgct tatatagacc tcccaccgta cacgcctacc gcccatttgc gtcaatgggg
     5221 cggagttgtt acgacatttt ggaaagtccc gttgattttg gtgccaaaac aaactcccat
     5281 tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc aaaccgctat ccacgcccat
     5341 tgatgtactg ccaaaaccgc atcaccatgg taatagcgat gactaatacg tagatgtact
     5401 gccaagtagg aaagtcccat aaggtcatgt actgggcata atgccaggcg ggccatttac
     5461 cgtcattgac gtcaataggg ggcgtacttg gcatatgata cacttgatgt actgccaagt
     5521 gggcagttta ccgtaaatac tccacccatt gacgtcaatg gaaagtccct attggcgtta
     5581 ctatgggaac atacgtcatt attgacgtca atgggcgggg gtcgttgggc ggtcagccag
     5641 gcgggccatt taccgtaagt tatgtaacgc ggaactccat atatgggcta tgaactaatg
     5701 accccgtaat tgattactat taataactaa tgcatggcgg taatacggtt atccacagaa
     5761 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5821 aaaaaggccg cgttgctggc gtt
//
