LOCUS       sequence-426474-        4072 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mAID-BromoTag for N-terminal tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4072)
  AUTHORS   Hatoyama Y, Islam M, Bond AG, Hayashi KI, Ciulli A, Kanemaki MT
  TITLE     Combination of AID2 and BromoTag expands the utility of degron-based
            protein knockdowns.
  JOURNAL   EMBO Rep. 2024 Sep;25(9):4062-4077. doi: 10.1038/s44319-024-00224-4.
            Epub 2024 Aug 23.
  PUBMED    39179892
REFERENCE   2  (bases 1 to 4072)
  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/s44319-024-00224-4"; journalName: "EMBO Rep"; date: 
            "2024-09"; volume: "25"; issue: "9"; pages: "4062-4077"
COMMENT     Sequence Label: pMK463 (BSD-P2A-mAB)
FEATURES             Location/Qualifiers
     source          1..4072
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      4..460
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(91..110)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     301..322
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     586..608
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     600..617
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     601..617
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     627..646
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        627..645
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     670..686
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     671..687
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      702..711
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             708..1103
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             1116..1172
                     /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             1179..1382
                     /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(1831..1847)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1831..1847)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(1884..1904)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1884..1902)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1923..1939)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1923..1939)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1936..1958)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1947..1963
                     /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(1971..1977,1978..1995,1996..2001))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2016..2037
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2154..2171)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2325..2913)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2405..2424)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3084..3875,3876..3944))
                     /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     3707..3726
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3945..4049)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
       61 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
      121 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
      181 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
      241 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
      301 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
      361 tataagggat tttgccgatt tcggcctatt ggttaaaaaa atgagctgat ttaacaaaaa
      421 tttaacgcga attttaacaa aatattaacg cttacaattt ccattcgcca ttcaggctgc
      481 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
      541 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
      601 gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa ttgggtaccg
      661 ggccccccct cgaggtcgac ggtatcgata agcttgatat cgccaccatg gccaagcctt
      721 tgtctcaaga agaatccacc ctcattgaaa gagcaacggc tacaatcaac agcatcccca
      781 tctctgaaga ctacagcgtc gccagcgcag ctctctctag cgacggccgc atcttcactg
      841 gtgtcaatgt atatcatttt actgggggac cttgtgcaga actcgtggtg ctgggcactg
      901 ctgctgctgc ggcagctggc aacctgactt gtatcgtcgc gatcggaaat gagaacaggg
      961 gcatcttgag cccctgcgga cggtgccgac aggtgcttct cgatctgcat cctgggatca
     1021 aagccatagt gaaggacagt gatggacagc cgacggcagt tgggattcgt gaattgctgc
     1081 cctctggtta tgtgtgggag ggctcaggaa gcggagctac taacttcagc ctgctgaagc
     1141 aggctggaga cgtggaggag aaccctggac ctagatctaa ggagaagagt gcttgtccta
     1201 aagatccagc caaacctccg gccaaggcac aagttgtggg atggccaccg gtgagatcat
     1261 accggaagaa cgtgatggtt tcctgccaaa aatcaagcgg tggcccggag gcggcggcgt
     1321 tcgtgaaggt atcaatggac ggagcaccgt acttgaggaa aatcgatttg aggatgtata
     1381 aaggcgctgg tgcaggcgcc ggatcagtga aggacgtgcc cgactctcag cagcacccag
     1441 caccagagaa gagcagcaag gtctcggagc agctcaagtg ctgcagcggc atcctcaagg
     1501 agatgtttgc caagaagcac gccgcctacg cctggccctt ctacaagcct gtggacgtgg
     1561 aggcactggg cgcccacgac tactgtgaca tcatcaagca ccccatggac atgagcacaa
     1621 tcaagtctaa actggaggcc cgtgagtacc gtgatgctca ggagtttggt gctgacgtcc
     1681 gattgatgtt ctccaactgc tataagtaca accctcctga ccatgaggtg gtggccatgg
     1741 cccgcaagct ccaggatgtg ttcgaaatgc gctttgccaa gatgccggac gagggcggag
     1801 gcggaggtgg cggcgctggt gcaggcgccg gatccactag ttctagagcg gccgccaccg
     1861 cggtggagct ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
     1921 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     1981 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     2041 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     2101 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     2161 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     2221 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     2281 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     2341 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     2401 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2461 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     2521 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2581 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2641 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     2701 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2761 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2821 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     2881 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     2941 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     3001 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     3061 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     3121 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     3181 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     3241 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     3301 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     3361 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     3421 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     3481 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     3541 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     3601 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     3661 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     3721 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     3781 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     3841 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     3901 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     3961 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     4021 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc ac
//
