LOCUS       sequence-442602-        7838 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of Myc-tagged BirA*-RNF8 (R42A) fusion protein 
            for BioID.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7838)
  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 7838)
  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: pcDNA3.1-Myc-BirA*-RNF8-R42A
FEATURES             Location/Qualifiers
     source          1..7838
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(12..31)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      825..1413
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1314..1333
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1567..1584
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1701..1722
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1737..1742,1743..1760,1761..1767)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1775..1791
                     /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     1780..1802
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1799..1815
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1799..1815
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1864..1985
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1875..1894)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1929..1948
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(2159..2953)
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2270..2289)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2880..2899
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(3020..3349)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3034..3169
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3088..3107)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     3334..3354
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(3363..3791)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3473..3494)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3685..3704
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    3837..4061
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     4050..4067
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(join(5596..6204,6205..6207,6208..6555))
                     /codon_start=1
                     /product="promiscuous R118G mutant of the E. coli biotin 
                     protein ligase (Cronan, 2005)"
                     /label=BirA(R118G)
                     /note="also known as BirA* or BioID"
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGGRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             complement(6559..6588)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     regulatory      6588..6597
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(6614..6633)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6615..6633)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(6678..6881)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(6707..6727)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        6882..7261
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     7433..7452
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        7527..7631
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7632..7700,7701..7838,1..654)
                     /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"
ORIGIN
        1 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
       61 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
      121 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
      181 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
      241 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
      301 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
      361 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
      421 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
      481 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
      541 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
      601 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
      661 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
      721 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
      781 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
      841 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
      901 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
      961 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     1021 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     1081 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     1141 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     1201 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     1261 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     1321 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     1381 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     1441 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     1501 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     1561 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc
     1621 cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg
     1681 ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta
     1741 cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca
     1801 ggaaacagct atgaccatga ttacgccaag ctctagctag aggtcgacgg tatacagaca
     1861 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     1921 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     1981 aagttggggt gggcgaagaa ctccagcatg agatccccgc gctggaggat catccagccg
     2041 gcgtcccgga aaacgattcc gaagcccaac ctttcataga aggcggcggt ggaatcgaaa
     2101 tctcgtgatg gcaggttggg cgtcgcttgg tcggtcattt cgaaccccag agtcccgctc
     2161 agaagaactc gtcaagaagg cgatagaagg cgatgcgctg cgaatcggga gcggcgatac
     2221 cgtaaagcac gaggaagcgg tcagcccatt cgccgccaag ctcttcagca atatcacggg
     2281 tagccaacgc tatgtcctga tagcggtccg ccacacccag ccggccacag tcgatgaatc
     2341 cagaaaagcg gccattttcc accatgatat tcggcaagca ggcatcgcca tgggtcacga
     2401 cgagatcctc gccgtcgggc atgcgcgcct tgagcctggc gaacagttcg gctggcgcga
     2461 gcccctgatg ctcttcgtcc agatcatcct gatcgacaag accggcttcc atccgagtac
     2521 gtgctcgctc gatgcgatgt ttcgcttggt ggtcgaatgg gcaggtagcc ggatcaagcg
     2581 tatgcagccg ccgcattgca tcagccatga tggatacttt ctcggcagga gcaaggtgag
     2641 atgacaggag atcctgcccc ggcacttcgc ccaatagcag ccagtccctt cccgcttcag
     2701 tgacaacgtc gagcacagct gcgcaaggaa cgcccgtcgt ggccagccac gatagccgcg
     2761 ctgcctcgtc ctgcagttca ttcagggcac cggacaggtc ggtcttgaca aaaagaaccg
     2821 ggcgcccctg cgctgacagc cggaacacgg cggcatcaga gcagccgatt gtctgttgtg
     2881 cccagtcata gccgaatagc ctctccaccc aagcggccgg agaacctgcg tgcaatccat
     2941 cttgttcaat catgcgaaac gatcctcatc ctgtctcttg atcagatccg aaaatggata
     3001 tacaagctcc cgggagcttt ttgcaaaagc ctaggcctcc aaaaaagcct cctcactact
     3061 tctggaatag ctcagaggca gaggcggcct cggcctctgc ataaataaaa aaaattagtc
     3121 agccatgggg cggagaatgg gcggaactgg gcggagttag gggcgggatg ggcggagtta
     3181 ggggcgggac tatggttgct gactaattga gatgcatgct ttgcatactt ctgcctgctg
     3241 gggagcctgg ggactttcca cacctggttg ctgactaatt gagatgcatg ctttgcatac
     3301 ttctgcctgc tggggagcct ggggactttc cacaccctaa ctgacacaca ttccacagaa
     3361 ttaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     3421 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     3481 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     3541 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     3601 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     3661 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     3721 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     3781 acagggcgcg tggggatacc ccctagagcc ccagctggtt ctttccgcct cagaagccat
     3841 agagcccacc gcatccccag catgcctgct attgtcttcc caatcctccc ccttgctgtc
     3901 ctgccccacc ccacccccca gaatagaatg acacctactc agacaatgcg atgcaatttc
     3961 ctcattttat taggaaagga cagtgggagt ggcaccttcc agggtcaagg aaggcacggg
     4021 ggaggggcaa acaacagatg gctggcaact agaaggcaca gtcgaggctg atcagcggtt
     4081 taaacttaag cttggtaccg agctcggatc cactagtcca gtgtggtgga attctcagaa
     4141 caatctcttt gcttttcgtt ccctaatgag aacaattcgt cgttctttca cttctgagct
     4201 cagattattt accatcttat taatgcaatt gtccagaacc aaagagtatg ttttggactt
     4261 aatgtccttc cgacaaatgg ggcattctat cttccgcttc atccattcat tgatacagta
     4321 ggagcagaaa ctgtgggcac agttcaaggt gacagcctca atgaagtatt ctgaacaaat
     4381 aatacattgg agctcattct ctagcacatc attcatgtgg ctaagaactt cttccttctg
     4441 tgcttgcatc ttctccttct cttccttggt ctgctctaat tctttgttct tggcttgaat
     4501 gattgcttca aagtccttct tgctgcgatt tagctcttcc attagagccc aatgctcctg
     4561 cagagcctgg gccagctgtt gcttcaggtc cttttctcct tgggctatct ccaaaccctg
     4621 aagatgctgt tcctcttcct ggaaagtctt ctctagttgc tccactcttg cctgctgctg
     4681 agcctgctct gcacacagct gggactgtaa gtcctgaagt tcctgctcca tttgcacaac
     4741 tttctttgag ttcccctttt gggtctgctt tttcacattc ataacggctt catgtttttc
     4801 ctgcatctgt attttgagcc tcagaatcct ggacatggtc accttaaaca tctgtaagct
     4861 tctctgagat gctgaagagt ttgaggcttt ctgctcatga tgaacctctg tgactttggg
     4921 gaagccaggg taaatgggag cccctgtggt cttacttggc tcaagggcag tcaacttagg
     4981 atccaaattg tcagagggtg tactggccac ttcacctttc ccctgtgatt tcactggctg
     5041 accagattca caagacactt tatttatttt ggatttcaaa tttgaggggc cttcagctcc
     5101 aggacctgct aattcatcca aactgaattt ccttttagtt ctcaattcct tatttttttc
     5161 tatcatttgg tcattctttg gggaaagaca aggatatatt gtctcccagt cttcttcagt
     5221 aacttcatat tcatactccg cattctcctt attttccaga ggcactccaa gttggatgta
     5281 gtctccctga tgaatggaat agacccttaa aggttccaga cgcgctctgt tcagccaaac
     5341 accatttaga ctcttgttgt ccataattgt ccattggccc tcaggattct gcttcaaaac
     5401 acagtggttt cgagaaatca tcagggggca gatttttgat accagttggt atgtgacacc
     5461 aaatcctgct cctacagtca cctcgcaccc atcttccagc agcagccacc cggcgctcat
     5521 ccccacccgc cgcaggcacc agctccggcc accggcgcgg tctcctgtga cgaagaagcc
     5581 gggctccccc tcgagcttct ctgcgcttct cagggagatt tctccgccca tccagggctt
     5641 gatgattccg tcctgctcca gcagcagagc tccctgcttg tcgattcctc tggagatgcc
     5701 gaagatttcc ttgtcgccga tgatcagctt cacaggtctg ttgatgaagt tgtccagctt
     5761 ctcccatctg ctcaggtaag gagccagtcc ctcctgctcg aacagctcca gagcggctct
     5821 cagctctctg atcagcatgg cggccagggt gttccggtcc aggttgatgc cagcctcctg
     5881 cagggtgatc cagccctggt tcaccacgct ctcctccact cttctcatgg ccatgttgat
     5941 tccggctccg atcacgatct gagcggcatc gcctgtcttg cctgtcagct ccaccaggat
     6001 gccagccagc ttgcggtcct ggaggtacag gtcattaggc cacttcactc tcaccttgtc
     6061 ggctcccagc tttctcagca cctcggccat cacgattccg atcaccagac tcagtccgat
     6121 ggcagcagca ggtccctgct ccagtctcca gaacatggac aggtacaggt tggctccgaa
     6181 tgggctgaac cacttccgtc ctctgcctcc tctgccagcc tgctggtact cggcgatgca
     6241 ggcgtcgccg gacttcagct ctccgattct gtccagcagg tactgattgg tggagtcgat
     6301 cacaggcagc acggccacgc ttccgccatc cagctgtccc aggatctgct tggcgttcag
     6361 cagctggata ggctcaggca ggctgtagcc cttgccaggc acggtgaaca cgtccactcc
     6421 ccagtctctc agtgtctgga tgtgcttgtt gatggcggct ctgctcattc ccagggtctc
     6481 tcccagctgc tcgccagagt ggaactcgcc gttggccagc agggcgatca gcttcagggg
     6541 cacggtgttg tccttgtcga gatcctcttc tgagatgagt ttttgttcca tggtggctag
     6601 ccagcttggg tctccctata gtgagtcgta ttaatttcga taagccagta agcagtgggt
     6661 tctctagtta gccagagagc tctgcttata tagacctccc accgtacacg cctaccgccc
     6721 atttgcgtca atggggcgga gttgttacga cattttggaa agtcccgttg attttggtgc
     6781 caaaacaaac tcccattgac gtcaatgggg tggagacttg gaaatccccg tgagtcaaac
     6841 cgctatccac gcccattgat gtactgccaa aaccgcatca ccatggtaat agcgatgact
     6901 aatacgtaga tgtactgcca agtaggaaag tcccataagg tcatgtactg ggcataatgc
     6961 caggcgggcc atttaccgtc attgacgtca atagggggcg tacttggcat atgatacact
     7021 tgatgtactg ccaagtgggc agtttaccgt aaatactcca cccattgacg tcaatggaaa
     7081 gtccctattg gcgttactat gggaacatac gtcattattg acgtcaatgg gcgggggtcg
     7141 ttgggcggtc agccaggcgg gccatttacc gtaagttatg taacgcggaa ctccatatat
     7201 gggctatgaa ctaatgaccc cgtaattgat tactattaat aactagtcaa taatcaatgt
     7261 caacgcgtat atctggcccg tacatcgcga agcagcgcaa aacgcctaac cctaagcaga
     7321 ttcttcatgc aattgtcggt caagccttgc cttgttgtag cttaaatttt gctcgcgcac
     7381 tactcagcga cctccaacac acaagcaggg agcagatact ggcttaacta tgcggcatca
     7441 gagcagattg tactgagagt gcaccatagg ggatcgggag atctcccgat ccgtcgacgt
     7501 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     7561 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     7621 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     7681 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     7741 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     7801 gttttcgccc cgaagaacgt tttccaatga tgagcact
//
