LOCUS       sequence-138172-        1058 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  C-terminal tagging with Stag-3FLAG using CRIPSR/Cas9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1058)
  AUTHORS   Natsume T, Kiyomitsu T, Saga Y, Kanemaki MT
  TITLE     Rapid Protein Depletion in Human Cells by Auxin-Inducible Degron 
            Tagging with Short Homology Donors.
  JOURNAL   Cell Rep. 2016 Apr 5;15(1):210-8. doi: 10.1016/j.celrep.2016.03.001.
            Epub 2016 Mar 24.
  PUBMED    27052166
REFERENCE   2  (bases 1 to 1058)
  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: 
            "10.1016/j.celrep.2016.03.001"; journalName: "Cell Rep"; date: 
            "2016-04-5- 5"; volume: "15"; issue: "1"; pages: "210-8"
COMMENT     Sequence Label: pMK284 (Stag-3FLAG-Hygro)
FEATURES             Location/Qualifiers
     source          1..1058
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     7..26
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7..25
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             58..102
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             115..180
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    312..433
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(349..368)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     403..422
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    464..497
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        558..1057
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(593..616)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     990..1009
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
ORIGIN
        1 cgcgcgtaat acgactcact atagggcgaa ttggagctcg gatccggtgc aggcgccaag
       61 gaaaccgccg ctgcaaaatt cgaacgccag cacatggaca gccgatcccg ggctgactac
      121 aaagaccatg acggtgatta taaagatcat gacatcgact acaaggatga cgatgacaag
      181 tagtgtacaa gtaatctaga taactgatca taatcagcca taccacattt gtagaggttt
      241 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
      301 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
      361 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
      421 tcaatgtatc ttaacgcgtc gatcatattc aataaccctt aatataactt cgtataatgt
      481 atgctatacg aagttattag gtctgaagag gagtttacgt ccagccaagc ttaggatctc
      541 gacctcgaaa ttctaccggg taggggaggc gcttttccca aggcagtctg gagcatgcgc
      601 tttagcagcc ccgctgggca cttggcgcta cacaagtggc ctctggcctc gcacacattc
      661 cacatccacc ggtaggcgcc aaccgactcc gttctttggt ggccccttcg cgccaccttc
      721 tactcctccc ctagtcagga agttcccccc cgccccgcag ctcgcgtcgt gcaggacgtg
      781 acaaatggaa gtagcacgtc tcactagtct cgtgcagatg gacagcaccg ctgagcaatg
      841 gaagcgggta ggcctttggg gcagcggcca atagcagctt tgctccttcg ctttctgggc
      901 tcagaggctg ggaaggggtg ggtccggggg cgggctcagg ggcgggctca ggggcggggc
      961 gggcgcccga aggtcctccg gaggcccggc attctgcacg cttcaaaagc gcacgtctgc
     1021 cgcgctgttc tcctcttcct catctccggg cctttcga
//
