LOCUS       sequence-234563-        4951 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  C-terminal tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4951)
  AUTHORS   Yesbolatova A, Natsume T, Hayashi KI, Kanemaki MT
  TITLE     Generation of conditional auxin-inducible degron (AID) cells and 
            tight control of degron-fused proteins using the degradation 
            inhibitor auxinole.
  JOURNAL   Methods. 2019 Apr 23. pii: S1046-2023(18)30331-1. doi: 
            10.1016/j.ymeth.2019.04.010.
  PUBMED    31026591
REFERENCE   2  (bases 1 to 4951)
  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: "Methods. 
            2019 Apr 23. pii: S1046-2023(18)30331-1. doi: 
            10.1016/j.ymeth.2019.04.010."
COMMENT     Sequence Label: pMK391 (mAID-BSD)
FEATURES             Location/Qualifiers
     source          1..4951
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     72..94
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     86..103
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     87..103
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     113..132
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        113..131
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             168..371
                     /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"
     polyA_signal    492..613
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(529..548)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     583..602
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    644..677
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        738..1237
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(773..796)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1170..1189
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      1261..1270
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1267..1665
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     protein_bind    2168..2201
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(2250..2270)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2250..2268)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2289..2305)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2289..2305)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2302..2324)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2313..2329
                     /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(2337..2343,2344..2361,2362..2367))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2382..2403
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2520..2537)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2691..3279)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2771..2790)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3450..4241,4242..4310))
                     /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     4073..4092
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4311..4415)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4442..4897
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4529..4548)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4739..4760
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
       61 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgagcgcg cgtaatacga
      121 ctcactatag ggcgaattgg agctccccgg gatccggtgc aggcgccaag gagaagagtg
      181 cttgtcctaa agatccagcc aaacctccgg ccaaggcaca agttgtggga tggccaccgg
      241 tgagatcata ccggaagaac gtgatggttt cctgccaaaa atcaagcggt ggcccggagg
      301 cggcggcgtt cgtgaaggta tcaatggacg gagcaccgta cttgaggaaa atcgatttga
      361 ggatgtataa ataatctaga taactgatca taatcagcca taccacattt gtagaggttt
      421 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
      481 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
      541 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
      601 tcaatgtatc ttaacgcgtc gatcatattc aataaccctt aatataactt cgtataatgt
      661 atgctatacg aagttattag gtctgaagag gagtttacgt ccagccaagc ttaggatctc
      721 gacctcgaaa ttctaccggg taggggaggc gcttttccca aggcagtctg gagcatgcgc
      781 tttagcagcc ccgctgggca cttggcgcta cacaagtggc ctctggcctc gcacacattc
      841 cacatccacc ggtaggcgcc aaccgactcc gttctttggt ggccccttcg cgccaccttc
      901 tactcctccc ctagtcagga agttcccccc cgccccgcag ctcgcgtcgt gcaggacgtg
      961 acaaatggaa gtagcacgtc tcactagtct cgtgcagatg gacagcaccg ctgagcaatg
     1021 gaagcgggta ggcctttggg gcagcggcca atagcagctt tgctccttcg ctttctgggc
     1081 tcagaggctg ggaaggggtg ggtccggggg cgggctcagg ggcgggctca ggggcggggc
     1141 gggcgcccga aggtcctccg gaggcccggc attctgcacg cttcaaaagc gcacgtctgc
     1201 cgcgctgttc tcctcttcct catctccggg cctttcgacc tgcatccatc tagatctcga
     1261 gccaccatgg ccaagccttt gtctcaagaa gaatccaccc tcattgaaag agcaacggct
     1321 acaatcaaca gcatccccat ctctgaagac tacagcgtcg ccagcgcagc tctctctagc
     1381 gacggccgca tcttcactgg tgtcaatgta tatcatttta ctgggggacc ttgtgcagaa
     1441 ctcgtggtgc tgggcactgc tgctgctgcg gcagctggca acctgacttg tatcgtcgcg
     1501 atcggaaatg agaacagggg catcttgagc ccctgcggac ggtgccgaca ggtgcttctc
     1561 gatctgcatc ctgggatcaa agccatagtg aaggacagtg atggacagcc gacggcagtt
     1621 gggattcgtg aattgctgcc ctctggttat gtgtgggagg gctaatcgat gatatcagat
     1681 ccccgggatg cagaaattga tgatctatta aacaataaag atgtccacta aaatggaagt
     1741 ttttcctgtc atactttgtt aagaagggtg agaacagagt acctacattt tgaatggaag
     1801 gattggagct acgggggtgg gggtggggtg ggattagata aatgcctgct ctttactgaa
     1861 ggctctttac tattgcttta tgataatgtt tcatagttgg atatcataat ttaaacaagc
     1921 aaaaccaaat taagggccag ctcattcctc ccactcatga tctatagatc tatagatctc
     1981 tcgtgggatc attgtttttc tcttgattcc cactttgtgg ttctaagtac tgtggtttcc
     2041 aaatgtgtca gtttcatagc ctgaagaacg agatcagcag cctctgttcc acatacactt
     2101 cattctcagt attgttttgc caagttctaa ttccatcaga agctggtcga gatccggaac
     2161 ccttaatata acttcgtata atgtatgcta tacgaagtta ttaggtccct cgaagaggtt
     2221 cactaggatc cggtacccag cttttgttcc ctttagtgag ggttaattgc gcgcttggcg
     2281 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     2341 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     2401 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     2461 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     2521 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     2581 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2641 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     2701 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     2761 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     2821 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     2881 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2941 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3001 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3061 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3121 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3181 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3241 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3301 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3361 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3421 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3481 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3541 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3601 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3661 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3721 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     3781 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     3841 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     3901 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     3961 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4021 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4081 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4141 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4201 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4261 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4321 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4381 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4441 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     4501 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     4561 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     4621 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     4681 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     4741 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     4801 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     4861 aacgcgaatt ttaacaaaat attaacgctt acaatttcca ttcgccattc aggctgcgca
     4921 actgttggga agggcgatcg gtgcgggcct c
//
