LOCUS       sequence-296095-        5167 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BRD4 tagging with mAID.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5167)
  AUTHORS   Yesbolatova A, Saito Y, Kitamoto N, Makino-Itou H, Ajima R, Nakano 
            R, Nakaoka H, Fukui K, Gamo K, Tominari Y, Takeuchi H, Saga Y, 
            Hayashi KI, Kanemaki MT
  TITLE     The auxin-inducible degron 2 technology provides sharp degradation 
            control in yeast, mammalian cells, and mice.
  JOURNAL   Nat Commun. 2020 Nov 11;11(1):5701. doi: 10.1038/s41467-020-19532-z.
  PUBMED    33177522
REFERENCE   2  (bases 1 to 5167)
  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: "Nat 
            Commun."; date: "2020-11-11"; volume: "11(1):5701. doi"; pages: " 
            10.1038/s41467-020-19532-z"
COMMENT     Sequence Label: mAID-BRD4 donor
FEATURES             Location/Qualifiers
     source          1..5167
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(24..452)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(134..155)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     346..365
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     579..601
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     593..610
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     594..610
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     620..639
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        620..638
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1181..2203
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             1181..2203
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             2216..2272
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2279..2482
                     /codon_start=1
                     /product="minimal degron from the auxin-responsive IAA17 
                     protein of Arabidopsis thaliana (Kubota et al., 2013)"
                     /label=mini-AID
                     /note="acts as an auxin-inducible degron (AID) in the 
                     presence of TIR1"
                     /translation="KEKSACPKDPAKPPAKAQVVGWPPVRSYRKNVMVSCQKSSGGPEA
                     AAFVKVSMDGAPYLRKIDLRMYK"
     primer_bind     complement(2973..2993)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2973..2991)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3012..3028)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3012..3028)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3025..3047)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3036..3052
                     /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(3060..3066,3067..3084,3085..3090))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3105..3126
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3243..3260)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3414..4002)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3494..3513)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4173..4964,4965..5033))
                     /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     4796..4815
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5034..5138)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
       61 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
      121 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
      181 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
      241 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
      301 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
      361 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
      421 cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg ccattcaggc tgcgcaactg
      481 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
      541 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
      601 gacggccagt gagcgcgcgt aatacgactc actatagggc gaattggagc tccaaatgaa
      661 atgattgact ttgctcgatg ctttgtgtgt gataaatgca aggagtttta atgtaaatgt
      721 catatgtggg agcctgcaca tcatcccctt ccacacacag gcagcatctc ctaaaaactc
      781 acaggtggga aaagggatgt gcaggtttgt gagacgtggc ttcattgtac atccttcccc
      841 atgaccttca tagttcccag agcctttgct ggctcctagt gggctctcac cacctcactc
      901 tgcctcctct gttggtttgt ggtctcctcc ctgctggagg gtaagtcttt ggggagagga
      961 gacattggaa ccctttcagt caaagcatgg ggtggtccct ctggccaact tggctagcta
     1021 ccgccaggga actcagtcct tggagaccac agccagaggg acttgtcatt ctgagcggtg
     1081 cccatctgct gactgatatc tcacgggggc tcttctcttc ctttgtagag tgcctggtga
     1141 agaatgtgat gggatcacta gcaagcttga tatcgccacc atgaaaaagc ctgaactcac
     1201 cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac agcgtctccg acctgatgca
     1261 gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt
     1321 cctgcgggta aatagctgcg ccgatggttt ctacaaagat cgttatgttt atcggcactt
     1381 tgcatcggcc gcgctcccga ttccggaagt gcttgacatt ggggaattca gcgagagcct
     1441 gacctattgc atctcccgcc gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga
     1501 actgcccgct gttctgcagc cggtcgcgga ggccatggat gcgatcgctg cggccgatct
     1561 tagccagacg agcgggttcg gcccattcgg accgcaagga atcggtcaat acactacatg
     1621 gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga
     1681 cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga
     1741 ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga
     1801 caatggccgc ataacagcgg tcattgactg gagcgaggcg atgttcgggg attcccaata
     1861 cgaggtcgcc aacatcttct tctggaggcc gtggttggct tgtatggagc agcagacgcg
     1921 ctacttcgag cggaggcatc cggagcttgc aggatcgccg cggctccggg cgtatatgct
     1981 ccgcattggt cttgaccaac tctatcagag cttggttgac ggcaatttcg atgatgcagc
     2041 ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg tcgggcgtac
     2101 acaaatcgcc cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag tactcgccga
     2161 tagtggaaac cgacgcccca gcactcgtcc gagggcaaag gaatcaggaa gcggagctac
     2221 taacttcagc ctgctgaagc aggctggaga cgtggaggag aaccctggac ctagatctaa
     2281 ggagaagagt gcttgtccta aagatccagc caaacctccg gccaaggcac aagttgtggg
     2341 atggccaccg gtgagatcat accggaagaa cgtgatggtt tcctgccaaa aatcaagcgg
     2401 tggcccggag gcggcggcgt tcgtgaaggt atcaatggac ggagcaccgt acttgaggaa
     2461 aatcgatttg aggatgtata aaggcgctgg tgcaggcgcc ggatccatgt ctgcggagag
     2521 tggtcctggg acgagattga gaaatctgcc agtaatgggg gatggactag aaacttccca
     2581 aatgtctaca acacaggccc aggcccaacc ccagccagcc aacgcagcca gcaccaaccc
     2641 cccgccccca gagacctcca accctaacaa gcccaagagg cagaccaacc aactgcaata
     2701 cctgctcaga gtggtgctca agacactatg gaaacaccag tttgcatggc ctttccagca
     2761 gcctgtggat gccgtcaagc tgaacctccc tgtgagtagt gtgggcaggg cccagggtgg
     2821 ccgtcctgga ggtcgtctcc tgcatgtctc ctggtcaggg cagttcctgg ggggagggta
     2881 ggagatgcat ttggaatcat ggtgagagct gcagggccct tgagtgagtg gtttcagcag
     2941 ttctgatcag ttgaggtacc cagcttttgt tccctttagt gagggttaat tgcgcgcttg
     3001 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3061 aacatacgag ccgggagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     3121 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3181 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     3241 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3301 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     3361 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3421 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     3481 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3541 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     3601 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     3661 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     3721 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     3781 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     3841 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     3901 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     3961 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4021 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4081 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4141 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4201 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4261 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4321 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4381 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4441 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     4501 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     4561 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     4621 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     4681 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     4741 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     4801 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     4861 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     4921 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     4981 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5041 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     5101 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     5161 cctaaat
//
