LOCUS       sequence-380616-        4500 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For membrane insertion study; expresses Tse5-CT (1169-1229)/PhoA 
            (22-472)/LacZ (4-60)fusion with PelB signalpeptide fused at the 
            N-terminus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4500)
  AUTHORS   Gonzalez-Magana A, Altuna J, Queralt-Martin M, Largo E, Velazquez C,
            Montanchez I, Bernal P, Alcaraz A, Albesa-Jove D
  TITLE     The P. aeruginosa effector Tse5 forms membrane pores disrupting the 
            membrane potential of intoxicated bacteria.
  JOURNAL   Commun Biol. 2022 Nov 5;5(1):1189. doi: 10.1038/s42003-022-04140-y.
  PUBMED    36335275
REFERENCE   2  (bases 1 to 4500)
  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: "Commun 
            Biol."; date: "2022-11-5"; volume: "5(1):1189. doi"; pages: " 
            10.1038/s42003-022-04140-y"
COMMENT     Sequence Label: pKTop::sptse5-CT (1169-1229)
FEATURES             Location/Qualifiers
     source          1..4500
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     sig_peptide     23..88
                     /codon_start=1
                     /gene="pelB (fragment)"
                     /product="leader peptide for secretion"
                     /label=pelB signal sequence
                     /translation="MKYLLPTAAAGLLLLAAQPAMA"
     primer_bind     complement(1651..1668)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1651..1667)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1660..1682)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1826..1845)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1945..1967
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2285..3079)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2396..2415)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3006..3025
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      3475..4020
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     3921..3940
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4137..4154
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4391..4412
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4427..4432,4433..4450,4451..4457)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4465..4481
                     /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)."
     primer_bind     4470..4492
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     join(4489..4500,1..5)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(4489..4500,1..5)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 atgaccatga ttacgccaag ctatgaagta cctgctgccg accgcggcgg cgggtctgct
       61 gctgctggcg gcgcagccgg cgatggcgat gggcctgatc attccgctgg ttgtgatcgg
      121 tgcgtttgcg gcgcgtgcgg cgattggtgc ggcgctgggt gcgggcattg agctgggtat
      181 gcagaccggc aaacaagtgc tgggtcagat gaaggataac tgggacagcg atcgtgacct
      241 gaccgatatc aagtggaaat gcatcgacat taactggaaa gggagctcgg cccggacacc
      301 agaaatgcct gttctggaaa accgggctgc tcagggcgat attactgcac ccggcggtgc
      361 tcgccgttta acgggtgatc agactgccgc tctgcgtgat tctcttagcg ataaacctgc
      421 aaaaaatatt attttgctga ttggcgatgg gatgggggac tcggaaatta ctgccgcacg
      481 taattatgcc gaaggtgcgg gcggcttttt taaaggtata gatgccttac cgcttaccgg
      541 gcaatacact cactatgcgc tgaataaaaa aaccggcaaa ccggactacg tcaccgactc
      601 ggctgcatca gcaaccgcct ggtcaaccgg tgtcaaaacc tataacggcg cgctgggcgt
      661 cgatattcac gaaaaagatc acccaacgat tctggaaatg gcaaaagccg caggtctggc
      721 gaccggtaac gtttctaccg cagagttgca ggatgccacg cccgctgcgc tggtggcaca
      781 tgtgacctcg cgcaaatgct acggtccgag cgcgaccagt gaaaaatgtc cgggtaacgc
      841 tctggaaaaa ggcggaaaag gatcgattac cgaacagctg cttaacgctc gtgccgacgt
      901 tacgcttggc ggcggcgcaa aaacctttgc tgaaacggca accgctggtg aatggcaggg
      961 aaaaacgctg cgtgaacagg cacaggcgcg tggttatcag ttggtgagcg atgctgcctc
     1021 actgaattcg gtgacggaag cgaatcagca aaaacccctg cttggcctgt ttgctgacgg
     1081 caatatgcca gtgcgctggc taggaccgaa agcaacgtac catggcaata tcgataagcc
     1141 cgcagtcacc tgtacgccaa atccgcaacg taatgacagt gtaccaaccc tggcgcagat
     1201 gaccgacaaa gccattgaat tgttgagtaa aaatgagaaa ggctttttcc tgcaagttga
     1261 aggtgcgtca atcgataaac aggatcatgc tgcgaatcct tgtgggcaaa ttggcgagac
     1321 ggtcgatctc gatgaagccg tacaacgggc gctggaattc gctaaaaagg agggtaacac
     1381 gctggtcata gtcaccgctg atcacgccca cgccagccag attgttgcgc cggataccaa
     1441 agctccgggc ctcacccagg cgctaaatac caaagatggc gcagtgatgg tgatgagtta
     1501 cgggaactcc gaagaggatt cacaagaaca taccggcagt cagttgcgta ttgcggcgta
     1561 tggcccgcat gccgccaatg ttgttggact gaccgaccag accgatctct tctacaccat
     1621 gaaagccgct ctggggctga aaacggattc actggccgtc gttttacaac gtcgtgactg
     1681 ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg
     1741 gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg
     1801 cgaatggcgc taagtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     1861 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     1921 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     1981 caccgaaacg cgcgagacga aagggccgcc cttcccaaca gttgcgcagc ctgaatggcg
     2041 aatggcgctg atgtccggcg gtgcttttgc cgttacgcac caccccgtca gtagctgaac
     2101 aggagggaca gggggtgggc gaagaactcc agcatgagat ccccgcgctg gaggatcatc
     2161 cagccggcgt cccggaaaac gattccgaag cccaaccttt catagaaggc ggcggtggaa
     2221 tcgaaatctc gtgatggcag gttgggcgtc gcttggtcgg tcatttcgaa ccccagagtc
     2281 ccgctcagaa gaactcgtca agaaggcgat agaaggcgat gcgctgcgaa tcgggagcgg
     2341 cgataccgta aagcacgagg aagcggtcag cccattcgcc gccaagctct tcagcaatat
     2401 cacgggtagc caacgctatg tcctgatagc ggtccgccac acccagccgg ccacagtcga
     2461 tgaatccaga aaagcggcca ttttccacca tgatattcgg caagcaggca tcgccatggg
     2521 tcacgacgag atcctcgccg tcgggcatcc gcgccttgag cctggcgaac agttcggctg
     2581 gcgcgagccc ctgatgctct tcgtccagat catcctgatc gacaagaccg gcttccatcc
     2641 gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc gaatgggcag gtagccggat
     2701 caagcgtatg cagccgccgc attgcatcag ccatgatgga tactttctcg gcaggagcaa
     2761 ggtgagatga caggagatcc tgccccggca cttcgcccaa tagcagccag tcccttcccg
     2821 cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc cgtcgtggcc agccacgata
     2881 gccgcgctgc ctcgtcttgg agttcattca gggcaccgga caggtcggtc ttgacaaaaa
     2941 gaaccgggcg cccctgcgct gacagccgga acacggcggc atcagagcag ccgattgtct
     3001 gttgtgccca gtcatagccg aatagcctct ccacccaagc ggccggagaa cctgcgtgca
     3061 atccatcttg ttcaatcatg cgaaacgatc ctcatcctgt ctcttgatca gatcttgatc
     3121 ccctgcgcca tcagatcctt ggcggcaaga aagccatcca gtttactttg cagggcttcc
     3181 caaccttacc agagggcgcc ccagctggca attccggttc gcttgctgtc cataaaaccg
     3241 cccagtctag ctatcgccat gtaagcccac tgcaagctac ctgctttctc tttgcgcttg
     3301 cgttttccct tgtccagata gcccagtagc tgacattcat ccggggtcag caccgtttct
     3361 gcggactggc tttctacgtg ttccgcttcc tttagcagcc cttgcgccct gagtgcttgc
     3421 ggcagcgtga agctgtgcct agaaatattt tatctgatta ataagatgat cttcttgaga
     3481 tcgttttggt ctgcgcgtaa tctcttgctc tgaaaacgaa aaaaccgcct tgcagggcgg
     3541 tttttcgaag gttctctgag ctaccaactc tttgaaccga ggtaactggc ttggaggagc
     3601 gcagtcacca aaacttgtcc tttcagttta gccttaaccg gcgcatgact tcaagactaa
     3661 ctcctctaaa tcaattacca gtggctgctg ccagtggtgc ttttgcatgt ctttccgggt
     3721 tggactcaag acgatagtta ccggataagg cgcagcggtc ggactgaacg gggggttcgt
     3781 gcatacagtc cagcttggag cgaactgcct acccggaact gagtgtcagg cgtggaatga
     3841 gacaaacgcg gccataacag cggaatgaca ccggtaaacc gaaaggcagg aacaggagag
     3901 cgcacgaggg agccgccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3961 caccactgat ttgagcgtca gatttcgtga tgcttgtcag gggggcggag cctatggaaa
     4021 aacggctttg ccgcggccct ctcacttccc tgttaagtat cttcctggca tcttccagga
     4081 aatctccgcc ccgttcgtaa gccatttccg ctcgccgcag tcgaacgacc gagcgtagcg
     4141 agtcagtgag cgaggaagcg gaatatatcc tgtatcacat attctgctga cgcaccggtg
     4201 cagccttttt tctcctgcca catgaagcac ttcactgaca ccctcatcag tgccaacata
     4261 gtaagccagt atatacactc cgctagcgcc caatacgcaa accgcctctc cccgcgcgtt
     4321 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc
     4381 gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc
     4441 ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct
//
