LOCUS       sequence-237529-        5160 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Theophylline-inducible expression of periplasmic APEX2 from a 
            mycobacterial shuttle plasmid (integrating).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5160)
  AUTHORS   Ganapathy US, Bai L, Wei L, Eckartt KA, Lett CM, Previti ML, Carrico
            IS, Seeliger JC
  TITLE     Compartment-Specific Labeling of Bacterial Periplasmic Proteins by 
            Peroxidase-Mediated Biotinylation.
  JOURNAL   ACS Infect Dis. 2018 Apr 30. doi: 10.1021/acsinfecdis.8b00044.
  PUBMED    29708735
REFERENCE   2  (bases 1 to 5160)
  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: "ACS Infect 
            Dis. 2018 Apr 30. doi: 10.1021/acsinfecdis.8b00044."
COMMENT     Sequence Label: pMV306-ribo-Sec-APEX2
FEATURES             Location/Qualifiers
     source          1..5160
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      1..10
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             51..92
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             93..839
                     /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"
     terminator      909..955
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(2892..2911)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2903..3718
                     /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="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPHAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHNLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2976..2995)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4049..4637
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4538..4557
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccatggcggc catcgcgacc tttgcggcac cggtcgcgtt ggctggatcc ggcaagccca
       61 tccccaaccc cctgctgggc ctggacagca ccggaaagtc ttacccaact gtgagtgctg
      121 attaccagga cgccgttgag aaggcgaaga agaagctcag aggcttcatc gctgagaaga
      181 gatgcgctcc tctaatgctc cgtttggcat tccactctgc tggaaccttt gacaagggca
      241 cgaagaccgg tggacccttc ggaaccatca agcaccctgc cgaactggct cacagcgcta
      301 acaacggtct tgacatcgct gttaggcttt tggagccact caaggcggag ttccctattt
      361 tgagctacgc cgatttctac cagttggctg gcgttgttgc cgttgaggtc acgggtggac
      421 ctaaggttcc attccaccct ggaagagagg acaagcctga gccaccacca gagggtcgct
      481 tgcccgatcc cactaagggt tctgaccatt tgagagatgt gtttggcaaa gctatggggc
      541 ttactgacca agatatcgtt gctctatctg ggggtcacac tattggagct gcacacaagg
      601 agcgttctgg atttgagggt ccctggacct ctaatcctct tattttcgac aactcatact
      661 tcacggagtt gttgagtggt gagaaggaag gtctccttca gctaccttct gacaaggctc
      721 ttttgtctga ccctgtattc cgccctctcg ttgataaata tgcagcggac gaagatgcct
      781 tctttgctga ttacgctgag gctcaccaaa agctttccga gcttgggttt gctgatgcct
      841 aatgaagatg gagaccaagc ttatcgatgt cgacgtagtt aactagcgta cgatcgactg
      901 ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatgccat
      961 catggccgcg gtgatcagct agagccgtga acgacagggc gaacgccagc ccgccgacgg
     1021 cgagggttcc gaccgctgca actcccggtg caaccttgtc ccggtctatt ctcttcactg
     1081 caccagctcc aatctggtgt gaatgcccct cgtctgttcg cgcaggcggg gggctctatt
     1141 cgtttgtcag catcgaaagt agccagatca gggatgcgtt gcaaccgcgt atgcccaggt
     1201 cagaagagtc gcacaagagt tgcagacccc tggaaagaaa aatggccaga gggcgaaaac
     1261 accctctgac cagcggagcg ggcgacggga atcgaacccg cgtagctagt ttggaagaat
     1321 gggtgtctgc cgaccacata tatgggccgg tcaagatagg tttttacccc ctctcggctg
     1381 catcctctaa gtggaaagaa attgcaggtc gtagaagcgc gttgaagcct gagagttgca
     1441 caggagttgc aacccggtag ccttgttcac gacgagagga gacctagttg gcacgtcgcg
     1501 gatggggatc gctgaagact cagcgcagcg ggaggatcca agcctcatac gtcaacccgc
     1561 aggacggtgt gaggtactac gcgctgcaga cctacgacaa caagatggac gccgaagcct
     1621 ggctcgcggg cgagaagcgg ctcatcgaga tggagacctg gacccctcca caggaccggg
     1681 cgaagaaggc agccgccagc gccatcacgc tggaggagta cacccggaag tggctcgtgg
     1741 agcgcgacct cgcagacggc accagggatc tgtacagcgg gcacgcggag cgccgcatct
     1801 acccggtgct aggtgaagtg gcggtcacag agatgacgcc agctctggtg cgtgcgtggt
     1861 gggccgggat gggtaggaag cacccgactg cccgccggca tgcctacaac gtcctccggg
     1921 cggtgatgaa cacagcggtc gaggacaagc tgatcgcaga gaacccgtgc cggatcgagc
     1981 agaaggcagc cgatgagcgc gacgtagagg cgctgacgcc tgaggagctg gacatcgtcg
     2041 ccgctgagat cttcgtgcac taccggatcg cggcatacat cctggcgtgg acgagcctcc
     2101 ggttcggaga gctgatcgag cttcgccgca aggacatcgt ggacgacggc atgacgatga
     2161 agctccgggt gcgccgtggc gcttcccgcg tggggaacaa gatcgtcgtt ggcaacgcca
     2221 agaccgtccg gtcgaagcgt cctgtgacgg ttccgcctca cgtcgcggag atgatccgag
     2281 cgcacatgaa ggaccgtacg aagatgaaca agggccccga ggcattcctg gtgaccacga
     2341 cgcagggcaa ccggctgtcg aagtccgcgt tcaccaagtc gctgaagcgt ggctacgcca
     2401 agatcggtcg gccggaactc cgcatccacg acctccgcgc tgtcggcgct acgttcgccg
     2461 ctcaggcagg tgcgacgacc aaggagctga tggcccgtct cggtcacacg actcctagga
     2521 tggcgatgaa gtaccagatg gcgtctgagg cccgcgacga ggctatcgct gaggcgatgt
     2581 ccaagctggc caagacctcc tgaaacgcaa aaagcccccc tcccaaggac actgagtcct
     2641 aaagaggggg gtttcttgtc agtacgcgaa gaaccacgcc tggccgcgag cgccagcacc
     2701 gccgctctgt gcggagacct gggcaccagc cccgccgccg ccaggagcat tgccgttccc
     2761 gccagctagc cacctgacgt cggggggggg ggaaagccac gttgtgtctc aaaatctctg
     2821 atgttacatt gcacaagata aaaatatatc atcatgaaca ataaaactgt ctgcttacat
     2881 aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtctt gctcgaggcc
     2941 gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc gcgataatgt
     3001 cgggcaatca ggtgcgacaa tctatcgctt gtatgggaag ccccatgcgc cagagttgtt
     3061 tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg tcagactaaa
     3121 ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta ctcctgatga
     3181 tgcatggtta ctcaccactg cgatccccgg gaaaacagca ttccaggtat tagaagaata
     3241 tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc ggttgcattc
     3301 gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg ctcaggcgca
     3361 atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc gtaatggctg
     3421 gcctgttgaa caagtctgga aagaaatgca taatcttttg ccattctcac cggattcagt
     3481 cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga aattaatagg
     3541 ttgtattgat gttggacgag tcggaatcgc agaccgatac caggatcttg ccatcctatg
     3601 gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa aatatggtat
     3661 tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt ttttctaatc
     3721 agaattggtt aattggttgt aacactggca gagcattacg ctgacttgac gggacggcgg
     3781 ctttgttgaa taaatcgaac ttttgctgag ttgaaggatc agatcacgca tcttcccgac
     3841 aacgcagacc gttccgtggc aaagcaaaag ttcaaaatca ccaactggtc cacctacaac
     3901 aaagctctca ccaaccgtgg ctccctcact ttctggctgg atgatggggc gattcaggcc
     3961 tggtatgagt cagcaacacc ttcttcacga ggcagacctc actagttcca tgagcgtcag
     4021 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     4081 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     4141 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     4201 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     4261 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     4321 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     4381 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     4441 tatgagaaag cgccacgctt cccgagggga gaaaggcgga caggtatccg gtaagcggca
     4501 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     4561 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     4621 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     4681 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     4741 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc aacgcgtgcg
     4801 gccgcggtac cagatcttta aatctagagg tgaccacaac gcgcccgctt tgatcgggga
     4861 cgtctgcggc cgaccattta cgggtcttgt tgtcgttggc ggtcatgggc cgaacatact
     4921 cacccggatc ggagggccga ggacaaggtc gaacgagggg catgacccgg tgcggggctt
     4981 cttgcactcg gcataggcga gtgctaagaa taacgttggc actcgcgacc gggatacgac
     5041 tcactatagg taccggtgat accagcatcg tcttgatgcc cttggcagca ccctgctaag
     5101 gaggcaacaa gatgcatatg aagctcacca caatgatcaa gacggcagta gcggtcgtgg
//
