LOCUS       sequence-442601-        6578 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of Myc-tagged human RNF8 
            (R477A/E478A/R479A/K480A).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6578)
  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 6578)
  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: pcDNA4.1-Myc-RNF8-4A
FEATURES             Location/Qualifiers
     source          1..6578
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(41..57)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(41..57)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(54..76)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    65..81
                     /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)."
     promoter        complement(join(89..95,96..113,114..119))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    134..155
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(272..289)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(443..1031)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(523..542)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1202..1993,1994..2062))
                     /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     1825..1844
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2063..2167)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2242..2261)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2433..2812
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2813..3016
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2967..2987
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    3018..3036
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3039..3057
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     3067..3091
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4699..4728
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4744..4761
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4784..4801)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4790..5014
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5060..5488
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5147..5166)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5357..5378
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5497..5517)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5502..5831
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5682..5817
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5744..5763
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5879..5926
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5945..6319
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    6449..6570
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6486..6505)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6540..6559
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
       61 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
      121 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
      181 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
      241 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
      301 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
      361 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
      421 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
      481 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
      541 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
      601 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
      661 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
      721 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
      781 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
      841 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
      901 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
      961 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     1021 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     1081 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     1141 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     1201 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     1261 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     1321 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     1381 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     1441 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     1501 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     1561 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     1621 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     1681 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     1741 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     1801 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     1861 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     1921 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     1981 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     2041 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     2101 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     2161 ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga
     2221 tcccctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct
     2281 gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac
     2341 aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct
     2401 gcttcgcgat gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata
     2461 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
     2521 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
     2581 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
     2641 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc
     2701 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg
     2761 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
     2821 gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
     2881 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
     2941 atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt
     3001 ctatataagc agagctctcc ctatcagtga tagagatctc cctatcagtg atagagatcg
     3061 tcgacgagct cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga
     3121 cctccataga agacaccggg accgatccag cctccggact ctagcgttta aacttaagct
     3181 tggtaccgag ctcggatcca ctagtccagt gtggtggaat tcatggggga gcccggcttc
     3241 ttcgtcacag gagaccgcgc cggtggccgg agctggtgcc tgcggcgggt ggggatgagc
     3301 gccgggtggc tgctgctgga agatgggtgc gaggtgactg taggacgagg atttggtgtc
     3361 acataccaac tggtatcaaa aatctgcccc ctgatgattt ctcgaaacca ctgtgttttg
     3421 aagcagaatc ctgagggcca atggacaatt atggacaaca agagtctaaa tggtgtttgg
     3481 ctgaacagag cgcgtctgga acctttaagg gtctattcca ttcatcaggg agactacatc
     3541 caacttggag tgcctctgga aaataaggag aatgcggagt atgaatatga agttactgaa
     3601 gaagactggg agacaatata tccttgtctt tccccaaaga atgaccaaat gatagaaaaa
     3661 aataaggaat tgagaactaa aaggaaattc agtttggatg aattagcagg tcctggagct
     3721 gaaggcccct caaatttgaa atccaaaata aataaagtgt cttgtgaatc tggtcagcca
     3781 gtgaaatcac aggggaaagg tgaagtggcc agtacaccct ctgacaattt ggatcctaag
     3841 ttgactgccc ttgagccaag taagaccaca ggggctccca tttaccctgg cttccccaaa
     3901 gtcacagagg ttcatcatga gcagaaagcc tcaaactctt cagcatctca gagaagctta
     3961 cagatgttta aggtgaccat gtccaggatt ctgaggctca aaatacagat gcaggaaaaa
     4021 catgaagccg ttatgaatgt gaaaaagcag acccaaaagg ggaactcaaa gaaagttgtg
     4081 caaatggagc aggaacttca ggacttacag tcccagctgt gtgcagagca ggctcagcag
     4141 caggcaagag tggagcaact agagaagact ttccaggaag aggaacagca tcttcagggt
     4201 ttggagatag cccaaggaga aaaggacctg aagcaacagc tggcccaggc tctgcaggag
     4261 cattgggctc taatggaaga gctaaatcgc agcaagaagg actttgaagc aatcattcaa
     4321 gccaagaaca aagaattaga gcagaccaag gaagagaagg agaagatgca agcacagaag
     4381 gaagaagttc ttagccacat gaatgatgtg ctagagaatg agctccaatg tattatttgt
     4441 tcagaatact tcattgaggc tgtcaccttg aactgtgccc acagtttctg ctcctactgt
     4501 atcaatgaat ggatgaagcg gaagatagaa tgccccattt gtcggaagga cattaagtcc
     4561 aaaacatact ctttggttct ggacaattgc attaataaga tggtaaataa tctgagctca
     4621 gaagtgaaag aacgacgaat tgttctcatt gcggctgcag cagcaaagag attgttcctc
     4681 gagtctagag ggcccttcga acaaaaactc atctcagaag aggatctgaa tatgcatacc
     4741 ggtcatcatc accatcacca ttgagtttaa acccgctgat cagcctcgac tgtgccttct
     4801 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     4861 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     4921 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     4981 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     5041 ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     5101 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     5161 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     5221 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     5281 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     5341 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     5401 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     5461 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     5521 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     5581 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     5641 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     5701 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     5761 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     5821 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtgt
     5881 tgacaattaa tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta
     5941 aaccatggcc aagttgacca gtgccgttcc ggtgctcacc gcgcgcgacg tcgccggagc
     6001 ggtcgagttc tggaccgacc ggctcgggtt ctcccgggac ttcgtggagg acgacttcgc
     6061 cggtgtggtc cgggacgacg tgaccctgtt catcagcgcg gtccaggacc aggtggtgcc
     6121 ggacaacacc ctggcctggg tgtgggtgcg cggcctggac gagctgtacg ccgagtggtc
     6181 ggaggtcgtg tccacgaact tccgggacgc ctccgggccg gccatgaccg agatcggcga
     6241 gcagccgtgg gggcgggagt tcgccctgcg cgacccggcc ggcaactgcg tgcacttcgt
     6301 ggccgaggag caggactgac acgtgctacg agatttcgat tccaccgccg ccttctatga
     6361 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     6421 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
     6481 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     6541 tggtttgtcc aaactcatca atgtatctta tcatgtct
//
