LOCUS       sequence-210102-        3391 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Multisite gateway vector for 3' tagging with APEX2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3391)
  AUTHORS   Ariotti N, Rae J, Giles N, Martel N, Sierecki E, Gambin Y, Hall TE, 
            Parton RG
  TITLE     Ultrastructural localisation of protein interactions using 
            conditionally stable nanobodies.
  JOURNAL   PLoS Biol. 2018 Apr 5;16(4):e2005473. doi: 
            10.1371/journal.pbio.2005473. eCollection 2018 Apr.
  PUBMED    29621251
REFERENCE   2  (bases 1 to 3391)
  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: "PLoS 
            Biol."; date: "2018-04-5"; volume: "16(4):e2005473. doi"; pages: " 
            10.1371/journal.pbio.2005473. eCollection 2018 Apr"
COMMENT     Sequence Label: p3E-APEX2
FEATURES             Location/Qualifiers
     source          1..3391
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     97..114
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(273..300)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(392..478)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     527..549
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     541..558
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     542..558
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(629..752)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             755..1501
                     /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="GKSYPTVSADYQDAVEKAKKKLRGFIAEKRCAPLMLRLAFHSAGT
                     FDKGTKTGGPFGTIKHPAELAHSANNGLDIAVRLLEPLKAEFPILSYADFYQLAGVVAV
                     EVTGGPKVPFHPGREDKPEPPPEGRLPDPTKGSDHLRDVFGKAMGLTDQDIVALSGGHT
                     IGAAHKERSGFEGPWTSNPLIFDNSYFTELLSGEKEGLLQLPSDKALLSDPVFRPLVDK
                     YAADEDAFFADYAEAHQKLSELGFADA"
     protein_bind    complement(1505..1600)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1637..1656)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1638..1656)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1661..1677)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1661..1677)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1779..1798)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1790..2599
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1857..1876)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2746..3334
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3235..3254
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
       61 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
      121 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
      181 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatacgcgt
      241 accgctagcc aggaagagtt tgtagaaacg caaaaaggcc atccgtcagg atggccttct
      301 gcttagtttg atgcctggca gtttatggcg ggcgtcctgc ccgccaccct ccgggccgtt
      361 gcttcacaac gttcaaatcc gctcccggcg gatttgtcct actcaggaga gcgttcaccg
      421 acaaacaaca gataaaacga aaggcccagt cttccgactg agcctttcgt tttatttgat
      481 gcctggcagt tccctactct cgcgttaacg ctagcatgga tgttttccca gtcacgacgt
      541 tgtaaaacga cggccagtct taagctcggg ccctgcagct ctagagctcg aattctacag
      601 gtcactaata ccatctaagt agttggttca tagtgactgc atatgttgtg ttttacagta
      661 ttatgtagtc tgttttttat gcaaaatcta atttaatata ttgatattta tatcatttta
      721 cgtttctcgt tcaactttct tgtacaaagt ggctggaaag tcttacccaa ctgtgagtgc
      781 tgattaccag gacgccgttg agaaggcgaa gaagaagctc agaggcttca tcgctgagaa
      841 gagatgcgct cctctaatgc tccgtttggc attccactct gctggaacct ttgacaaggg
      901 cacgaagacc ggtggaccct tcggaaccat caagcaccct gccgaactgg ctcacagcgc
      961 taacaacggt cttgacatcg ctgttaggct tttggagcca ctcaaggcgg agttccctat
     1021 tttgagctac gccgatttct accagttggc tggcgttgtt gccgttgagg tcacgggtgg
     1081 acctaaggtt ccattccacc ctggaagaga ggacaagcct gagccaccac cagagggtcg
     1141 cttgcccgat cccactaagg gttctgacca tttgagagat gtgtttggca aagctatggg
     1201 gcttactgac caagatatcg ttgctctatc tgggggtcac actattggag ctgcacacaa
     1261 ggagcgttct ggatttgagg gtccctggac ctctaatcct cttattttcg acaactcata
     1321 cttcacggag ttgttgagtg gtgagaagga aggtctcctt cagctacctt ctgacaaggc
     1381 tcttttgtct gaccctgtat tccgccctct cgttgacaaa tatgcagcgg acgaagatgc
     1441 cttctttgct gattacgctg aggctcacca aaagctttcc gagcttgggt ttgctgatgc
     1501 ctaacaactt tattatacaa agttggcatt ataaaaaagc attgcttatc aatttgttgc
     1561 aacgaacagg tcactatcag tcaaaataaa atcattattt ggagctccat ggtagcgtta
     1621 acgcggccgc gatatcccct atagtgagtc gtattacatg gtcatagctg tttcctggca
     1681 gctctggccc gtgtctcaaa atctctgatg ttacattgca caagataaaa atatatcatc
     1741 atgaacaata aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat
     1801 tcaacgggaa acgtcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta
     1861 taaatgggct cgcgataatg tcgggcaatc aggtgcgaca atctatcgct tgtatgggaa
     1921 gcccgatgcg ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac
     1981 agatgagatg gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca
     2041 ttttatccgt actcctgatg atgcatggtt actcaccact gcgatccccg gaaaaacagc
     2101 attccaggta ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt
     2161 gttcctgcgc cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt
     2221 atttcgtctc gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt
     2281 tgatgacgag cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataaactttt
     2341 gccattctca ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt
     2401 tgacgagggg aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata
     2461 ccaggatctt gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg
     2521 gctttttcaa aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat
     2581 gctcgatgag tttttctaat cagaattggt taattggttg taacactggc agagcattac
     2641 gctgacttga cgggacggcg caagctcatg accaaaatcc cttaacgtga gttacgcgtc
     2701 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     2761 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     2821 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     2881 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     2941 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     3001 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     3061 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     3121 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     3181 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     3241 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     3301 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     3361 gttcctggcc ttttgctggc cttttgctca c
//
