LOCUS       sequence-439512-        5752 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of FLAG-tagged human RNF8.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5752)
  AUTHORS   Chuah YH, Tay EXY, Grinchuk OV, Yoon J, Feng J, Kannan S, Robert M, 
            Jakhar R, Liang Y, Lee BWL, Wang LC, Lim YT, Zhao T, Sobota RM, Lu 
            G, Low BC, Crasta KC, Verma CS, Lin Z, Ong DST
  TITLE     CAMK2D serves as a molecular scaffold for RNF8-MAD2 complex to 
            induce mitotic checkpoint in glioma.
  JOURNAL   Cell Death Differ. 2023 Aug;30(8):1973-1987. doi: 
            10.1038/s41418-023-01192-3. Epub 2023 Jul 19.
  PUBMED    37468549
REFERENCE   2  (bases 1 to 5752)
  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"; doi: 
            "10.1038/s41418-023-01192-3"; journalName: "Cell Death Differ"; 
            date: "2023-08"; volume: "30"; issue: "8"; pages: "1973-1987"
COMMENT     Sequence Label: pCMV-FLAG-RNF8
FEATURES             Location/Qualifiers
     source          1..5752
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        105..408
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        409..612
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     563..583
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        658..676
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      711..720
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             720..743
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2291..2310)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2292..2310)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    2584..2705
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2621..2640)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2675..2694
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2712..3167)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2849..2870)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3061..3080
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3194..3298
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3300..3657
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3300..3491
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3323..3343)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3508..3643
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3692..4486
                     /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             3692..4486
                     /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(3746..3765)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4356..4375
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4674..4693)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4718..4765
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5094..5682
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5583..5602
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttatcccc tgattctgtg gataaccgta ttaccgccat gcattagtta ttaatagtaa
       61 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg
      121 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
      181 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
      241 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
      301 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
      361 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt
      421 tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac
      481 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
      541 cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
      601 ataagcagag ctggtttagt gaaccgtcag atccgctagc gattacgcca agctcgaaat
      661 taaccctcac taaagggaac aaaagctgga gctccaccgc ggtggcggcc gccaccatgg
      721 attacaagga tgacgacgat aagagcccgg gcggatcccc cgggctgcag gaattcatgg
      781 gggagcccgg cttcttcgtc acaggagacc gcgccggtgg ccggagctgg tgcctgcggc
      841 gggtggggat gagcgccggg tggctgctgc tggaagatgg gtgcgaggtg actgtaggac
      901 gaggatttgg tgtcacatac caactggtat caaaaatctg ccccctgatg atttctcgaa
      961 accactgtgt tttgaagcag aatcctgagg gccaatggac aattatggac aacaagagtc
     1021 taaatggtgt ttggctgaac agagcgcgtc tggaaccttt aagggtctat tccattcatc
     1081 agggagacta catccaactt ggagtgcctc tggaaaataa ggagaatgcg gagtatgaat
     1141 atgaagttac tgaagaagac tgggagacaa tatatccttg tctttcccca aagaatgacc
     1201 aaatgataga aaaaaataag gaattgagaa ctaaaaggaa attcagtttg gatgaattag
     1261 caggtcctgg agctgaaggc ccctcaaatt tgaaatccaa aataaataaa gtgtcttgtg
     1321 aatctggtca gccagtgaaa tcacagggga aaggtgaagt ggccagtaca ccctctgaca
     1381 atttggatcc taagttgact gcccttgagc caagtaagac cacaggggct cccatttacc
     1441 ctggcttccc caaagtcaca gaggttcatc atgagcagaa agcctcaaac tcttcagcat
     1501 ctcagagaag cttacagatg tttaaggtga ccatgtccag gattctgagg ctcaaaatac
     1561 agatgcagga aaaacatgaa gccgttatga atgtgaaaaa gcagacccaa aaggggaact
     1621 caaagaaagt tgtgcaaatg gagcaggaac ttcaggactt acagtcccag ctgtgtgcag
     1681 agcaggctca gcagcaggca agagtggagc aactagagaa gactttccag gaagaggaac
     1741 agcatcttca gggtttggag atagcccaag gagaaaagga cctgaagcaa cagctggccc
     1801 aggctctgca ggagcattgg gctctaatgg aagagctaaa tcgcagcaag aaggactttg
     1861 aagcaatcat tcaagccaag aacaaagaat tagagcagac caaggaagag aaggagaaga
     1921 tgcaagcaca gaaggaagaa gttcttagcc acatgaatga tgtgctagag aatgagctcc
     1981 aatgtattat ttgttcagaa tacttcattg aggctgtcac cttgaactgt gcccacagtt
     2041 tctgctccta ctgtatcaat gaatggatga agcggaagat agaatgcccc atttgtcgga
     2101 aggacattaa gtccaaaaca tactctttgg ttctggacaa ttgcattaat aagatggtaa
     2161 ataatctgag ctcagaagtg aaagaacgac gaattgttct cattagggaa cgaaaagcaa
     2221 agagattgtt cctcgagggg gggcccggta ccttaattaa ttaaggtacc aggtaagtgt
     2281 acccaattcg ccctatagtg agtcgtatta caattcactc gatcgccctt cccaacagtt
     2341 gcgcagcctg aatggcgaat ggagatccaa tttttaagtg tataatgtgt taaactactg
     2401 attctaattg tttgtgtatt ttagattcac agtcccaagg ctcatttcag gcccctcagt
     2461 cctcacagtc tgttcatgat cataatcagc cataccacat ttgtagaggt tttacttgct
     2521 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt
     2581 gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     2641 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     2701 tcttaacgcg taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     2761 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     2821 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     2881 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     2941 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     3001 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     3061 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     3121 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt
     3181 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     3241 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagaatcc
     3301 tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     3361 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     3421 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     3481 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     3541 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     3601 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat
     3661 cgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     3721 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     3781 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     3841 gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct
     3901 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     3961 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     4021 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     4081 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     4141 cggtcttgtc gatcaggatg atctggacga agaacatcag gggctcgcgc cagccgaact
     4201 gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     4261 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     4321 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     4381 agaacttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     4441 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     4501 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc
     4561 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     4621 cagcgcgggg atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca
     4681 cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa
     4741 acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     4801 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     4861 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     4921 cctgccatag cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa
     4981 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     5041 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     5101 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     5161 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5221 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     5281 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     5341 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     5401 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     5461 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     5521 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     5581 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     5641 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     5701 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tg
//
