LOCUS       sequence-419271-        2151 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Cytosolic APEX2 control. Can be used as a C-terminal APEX2 fusion by
            digestion with NheI and Gibson cloning (see comments)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 2151)
  AUTHORS   Hollingsworth LR, Veeraraghavan P, Paulo JA, Harper JW
  TITLE     Spatiotemporal proteomic profiling of cellular responses to NLRP3 
            agonists
  JOURNAL   bioRxiv 2024.04.19.590338
REFERENCE   2  (bases 1 to 2151)
  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: "bioRxiv 
            2024.04.19.590338"
COMMENT     Sequence Label: pLV UBC APEX2-3xFLAG-IRES-eGFP
FEATURES             Location/Qualifiers
     source          1..2151
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             1..750
                     /codon_start=1
                     /product="soybean ascorbate peroxidase, engineered to serve
                     as a monomeric tag for electron microscopy and live-cell 
                     proteomics (Lam et al., 2015)"
                     /label=APEX2
                     /note="more active than the original APEX due to the A134P 
                     mutation"
                     /translation="MGKSYPTVSADYQDAVEKAKKKLRGFIAEKRCAPLMLRLAFHSAG
                     TFDKGTKTGGPFGTIKHPAELAHSANNGLDIAVRLLEPLKAEFPILSYADFYQLAGVVA
                     VEVTGGPKVPFHPGREDKPEPPPEGRLPDPTKGSDHLRDVFGKAMGLTDQDIVALSGGH
                     TIGAAHKERSGFEGPWTSNPLIFDNSYFTELLSGEKEGLLQLPSDKALLSDPVFRPLVD
                     KYAADEDAFFADYAEAHQKLSELGFADA"
     CDS             769..834
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     misc_feature    871..1423
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1015..1032)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1242..1261
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(1435..1437,1438..1440,1441..2151)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1435..1437,1438..1440,1441..2151)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1480..1501)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1741..1760)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2088..2109
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 atgggaaagt cttacccaac tgtgagtgct gattaccagg acgccgttga gaaggcgaag
       61 aagaagctca gaggcttcat cgctgagaag agatgcgctc ctctaatgct ccgtttggca
      121 ttccactctg ctggaacctt tgacaagggc acgaagaccg gtggaccctt cggaaccatc
      181 aagcaccctg ccgaactggc tcacagcgct aacaacggtc ttgacatcgc tgttaggctt
      241 ttggagccac tcaaggcgga gttccctatt ttgagctacg ccgatttcta ccagttggct
      301 ggcgttgttg ccgttgaggt cacgggtgga cctaaggttc cattccaccc tggaagagag
      361 gacaagcctg agccaccacc agagggtcgc ttgcccgatc ccactaaggg ttctgaccat
      421 ttgagagatg tgtttggcaa agctatgggg cttactgacc aagatatcgt tgctctatct
      481 gggggtcaca ctattggagc tgcacacaag gagcgttctg gatttgaggg tccctggacc
      541 tctaatcctc ttattttcga caactcatac ttcacggagt tgttgagtgg tgagaaggaa
      601 ggtctccttc agctaccttc tgacaaggct cttttgtctg accctgtatt ccgccctctc
      661 gttgacaaat atgcagcgga cgaagatgcc ttctttgctg attacgctga ggctcaccaa
      721 aagctttccg agcttgggtt tgctgatgcc ggcggatccg ggggaagcga ctacaaggac
      781 cacgacggtg actacaagga ccacgacatc gactacaagg acgacgacga caagtaatga
      841 gtcgacatta attaacgccc ccccccccta acgttactgg ccgaagccgc ttggaataag
      901 gccggtgtgc gtttgtctat atgttatttt ccaccatatt gccgtctttt ggcaatgtga
      961 gggcccggaa acctggccct gtcttcttga cgagcattcc taggggtctt tcccctctcg
     1021 ccaaaggaat gcaaggtctg ttgaatgtcg tgaaggaagt agttcctctg gaagcttctt
     1081 gaagacaaac aacgtctgta gcgacccttt gcaggcagcg gaacccccca cctggcgaca
     1141 ggtgcctctg cggccaaaag ccacgtgtat aagatacacc tgcaaaggcg gcacaacccc
     1201 agtgccacgt tgtgagttgg atagttgtgg aaagagtcaa atggctctcc tcaagcgtat
     1261 tcaacaaggg gctgaaggat gcccagaagg taccccattg tatgggatct gatctggggc
     1321 ctcggtgcac atgctttaca tgtatttagt cgaggttaaa aaaacgtcta ggccccccga
     1381 accacgggga cgtggttttc ctttgaaaaa cacgatgata atatggccac aaccatggtg
     1441 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     1501 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     1561 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     1621 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     1681 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     1741 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     1801 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     1861 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
     1921 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     1981 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     2041 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     2101 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa g
//
