LOCUS       sequence-230641-        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"
     rep_origin      4..459
                     /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     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             681..884
                     /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    1005..1126
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1042..1061)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1096..1115
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1157..1190
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1251..1750
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1286..1309)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1683..1702
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      1774..1783
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1780..2178
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     protein_bind    2681..2714
                     /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(2763..2783)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2763..2781)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2802..2818)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2802..2818)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2815..2837)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2826..2842
                     /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(2850..2856,2857..2874,2875..2880))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2895..2916
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3033..3050)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3204..3792)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3284..3303)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3963..4754,4755..4823))
                     /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     4586..4605
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4824..4928)
                     /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 tgagctgatt taacaaaaat
      421 ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggagctccc
      661 cgggatccgg tgcaggcgcc aaggagaaga gtgcttgtcc taaagatcca gccaaacctc
      721 cggccaaggc acaagttgtg ggatggccac cggtgagatc ataccggaag aacgtgatgg
      781 tttcctgcca aaaatcaagc ggtggcccgg aggcggcggc gttcgtgaag gtatcaatgg
      841 acggagcacc gtacttgagg aaaatcgatt tgaggatgta taaataatct agataactga
      901 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
      961 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     1021 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     1081 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc gtcgatcata
     1141 ttcaataacc cttaatataa cttcgtataa tgtatgctat acgaagttat taggtctgaa
     1201 gaggagttta cgtccagcca agcttaggat ctcgacctcg aaattctacc gggtagggga
     1261 ggcgcttttc ccaaggcagt ctggagcatg cgctttagca gccccgctgg gcacttggcg
     1321 ctacacaagt ggcctctggc ctcgcacaca ttccacatcc accggtaggc gccaaccgac
     1381 tccgttcttt ggtggcccct tcgcgccacc ttctactcct cccctagtca ggaagttccc
     1441 ccccgccccg cagctcgcgt cgtgcaggac gtgacaaatg gaagtagcac gtctcactag
     1501 tctcgtgcag atggacagca ccgctgagca atggaagcgg gtaggccttt ggggcagcgg
     1561 ccaatagcag ctttgctcct tcgctttctg ggctcagagg ctgggaaggg gtgggtccgg
     1621 gggcgggctc aggggcgggc tcaggggcgg ggcgggcgcc cgaaggtcct ccggaggccc
     1681 ggcattctgc acgcttcaaa agcgcacgtc tgccgcgctg ttctcctctt cctcatctcc
     1741 gggcctttcg acctgcatcc atctagatct cgagccacca tggccaagcc tttgtctcaa
     1801 gaagaatcca ccctcattga aagagcaacg gctacaatca acagcatccc catctctgaa
     1861 gactacagcg tcgccagcgc agctctctct agcgacggcc gcatcttcac tggtgtcaat
     1921 gtatatcatt ttactggggg accttgtgca gaactcgtgg tgctgggcac tgctgctgct
     1981 gcggcagctg gcaacctgac ttgtatcgtc gcgatcggaa atgagaacag gggcatcttg
     2041 agcccctgcg gacggtgccg acaggtgctt ctcgatctgc atcctgggat caaagccata
     2101 gtgaaggaca gtgatggaca gccgacggca gttgggattc gtgaattgct gccctctggt
     2161 tatgtgtggg agggctaatc gatgatatca gatccccggg atgcagaaat tgatgatcta
     2221 ttaaacaata aagatgtcca ctaaaatgga agtttttcct gtcatacttt gttaagaagg
     2281 gtgagaacag agtacctaca ttttgaatgg aaggattgga gctacggggg tgggggtggg
     2341 gtgggattag ataaatgcct gctctttact gaaggctctt tactattgct ttatgataat
     2401 gtttcatagt tggatatcat aatttaaaca agcaaaacca aattaagggc cagctcattc
     2461 ctcccactca tgatctatag atctatagat ctctcgtggg atcattgttt ttctcttgat
     2521 tcccactttg tggttctaag tactgtggtt tccaaatgtg tcagtttcat agcctgaaga
     2581 acgagatcag cagcctctgt tccacataca cttcattctc agtattgttt tgccaagttc
     2641 taattccatc agaagctggt cgagatccgg aacccttaat ataacttcgt ataatgtatg
     2701 ctatacgaag ttattaggtc cctcgaagag gttcactagg atccggtacc cagcttttgt
     2761 tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg
     2821 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     2881 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     2941 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     3001 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     3061 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     3121 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     3181 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     3241 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     3301 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     3361 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     3421 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3481 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3541 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     3601 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     3661 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     3721 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     3781 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     3841 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     3901 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     3961 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     4021 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     4081 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     4141 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     4201 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     4261 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     4321 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     4381 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     4441 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     4501 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     4561 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     4621 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     4681 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     4741 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     4801 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     4861 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     4921 gttccgcgca catttccccg aaaagtgcca c
//
