LOCUS       sequence-380613-        4692 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For membrane insertion study; Expresses tse5-CT (1169-1317) /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 4692)
  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 4692)
  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::tse5-CT (1169-1317)
FEATURES             Location/Qualifiers
     source          1..4692
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    110..131
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(146..151,152..169,170..176)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    184..200
                     /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     189..211
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     208..224
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     208..224
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2062..2079)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2062..2078)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2071..2093)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2237..2256)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2356..2378
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2696..3490)
                     /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(2807..2826)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3417..3436
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      3886..4431
                     /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     4332..4351
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4548..4565
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gctagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct
       61 ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt
      121 agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg
      181 gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccaagc
      241 tatgatcatt ccgctggttg tgatcggtgc gtttgcggcg cgtgcggcga ttggtgcggc
      301 gctgggtgcg ggcattgagc tgggtatgca gaccggcaaa caagtgctgg gtcagatgaa
      361 ggataactgg gacagcgatc gtgacctgac cgatatcaag tggaaatgca tcgacattaa
      421 ctggaaacat gttggtgcga gcgcggcgat tggtaccgtt gcgccgggca tgctgagcac
      481 cggtaaaacc gtggttcaga gcgcgaaggc gattcgtacc ctgagcggtc aggcggcgaa
      541 caccgcgaac cgtgcggcga aactggcggc gcgtaaggcg gcgcacgcgg ataccatcaa
      601 gaaagcggtg gcgacccaag cggcgtggca gaccggcaag caaattgtta aatgcccgct
      661 gaaggacgag gaagaggaat gcccgccgca agggagctcg gcccggacac cagaaatgcc
      721 tgttctggaa aaccgggctg ctcagggcga tattactgca cccggcggtg ctcgccgttt
      781 aacgggtgat cagactgccg ctctgcgtga ttctcttagc gataaacctg caaaaaatat
      841 tattttgctg attggcgatg ggatggggga ctcggaaatt actgccgcac gtaattatgc
      901 cgaaggtgcg ggcggctttt ttaaaggtat agatgcctta ccgcttaccg ggcaatacac
      961 tcactatgcg ctgaataaaa aaaccggcaa accggactac gtcaccgact cggctgcatc
     1021 agcaaccgcc tggtcaaccg gtgtcaaaac ctataacggc gcgctgggcg tcgatattca
     1081 cgaaaaagat cacccaacga ttctggaaat ggcaaaagcc gcaggtctgg cgaccggtaa
     1141 cgtttctacc gcagagttgc aggatgccac gcccgctgcg ctggtggcac atgtgacctc
     1201 gcgcaaatgc tacggtccga gcgcgaccag tgaaaaatgt ccgggtaacg ctctggaaaa
     1261 aggcggaaaa ggatcgatta ccgaacagct gcttaacgct cgtgccgacg ttacgcttgg
     1321 cggcggcgca aaaacctttg ctgaaacggc aaccgctggt gaatggcagg gaaaaacgct
     1381 gcgtgaacag gcacaggcgc gtggttatca gttggtgagc gatgctgcct cactgaattc
     1441 ggtgacggaa gcgaatcagc aaaaacccct gcttggcctg tttgctgacg gcaatatgcc
     1501 agtgcgctgg ctaggaccga aagcaacgta ccatggcaat atcgataagc ccgcagtcac
     1561 ctgtacgcca aatccgcaac gtaatgacag tgtaccaacc ctggcgcaga tgaccgacaa
     1621 agccattgaa ttgttgagta aaaatgagaa aggctttttc ctgcaagttg aaggtgcgtc
     1681 aatcgataaa caggatcatg ctgcgaatcc ttgtgggcaa attggcgaga cggtcgatct
     1741 cgatgaagcc gtacaacggg cgctggaatt cgctaaaaag gagggtaaca cgctggtcat
     1801 agtcaccgct gatcacgccc acgccagcca gattgttgcg ccggatacca aagctccggg
     1861 cctcacccag gcgctaaata ccaaagatgg cgcagtgatg gtgatgagtt acgggaactc
     1921 cgaagaggat tcacaagaac ataccggcag tcagttgcgt attgcggcgt atggcccgca
     1981 tgccgccaat gttgttggac tgaccgacca gaccgatctc ttctacacca tgaaagccgc
     2041 tctggggctg aaaacggatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc
     2101 tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag
     2161 cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggcg
     2221 ctaagtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     2281 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     2341 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     2401 gcgcgagacg aaagggccgc ccttcccaac agttgcgcag cctgaatggc gaatggcgct
     2461 gatgtccggc ggtgcttttg ccgttacgca ccaccccgtc agtagctgaa caggagggac
     2521 agggggtggg cgaagaactc cagcatgaga tccccgcgct ggaggatcat ccagccggcg
     2581 tcccggaaaa cgattccgaa gcccaacctt tcatagaagg cggcggtgga atcgaaatct
     2641 cgtgatggca ggttgggcgt cgcttggtcg gtcatttcga accccagagt cccgctcaga
     2701 agaactcgtc aagaaggcga tagaaggcga tgcgctgcga atcgggagcg gcgataccgt
     2761 aaagcacgag gaagcggtca gcccattcgc cgccaagctc ttcagcaata tcacgggtag
     2821 ccaacgctat gtcctgatag cggtccgcca cacccagccg gccacagtcg atgaatccag
     2881 aaaagcggcc attttccacc atgatattcg gcaagcaggc atcgccatgg gtcacgacga
     2941 gatcctcgcc gtcgggcatc cgcgccttga gcctggcgaa cagttcggct ggcgcgagcc
     3001 cctgatgctc ttcgtccaga tcatcctgat cgacaagacc ggcttccatc cgagtacgtg
     3061 ctcgctcgat gcgatgtttc gcttggtggt cgaatgggca ggtagccgga tcaagcgtat
     3121 gcagccgccg cattgcatca gccatgatgg atactttctc ggcaggagca aggtgagatg
     3181 acaggagatc ctgccccggc acttcgccca atagcagcca gtcccttccc gcttcagtga
     3241 caacgtcgag cacagctgcg caaggaacgc ccgtcgtggc cagccacgat agccgcgctg
     3301 cctcgtcttg gagttcattc agggcaccgg acaggtcggt cttgacaaaa agaaccgggc
     3361 gcccctgcgc tgacagccgg aacacggcgg catcagagca gccgattgtc tgttgtgccc
     3421 agtcatagcc gaatagcctc tccacccaag cggccggaga acctgcgtgc aatccatctt
     3481 gttcaatcat gcgaaacgat cctcatcctg tctcttgatc agatcttgat cccctgcgcc
     3541 atcagatcct tggcggcaag aaagccatcc agtttacttt gcagggcttc ccaaccttac
     3601 cagagggcgc cccagctggc aattccggtt cgcttgctgt ccataaaacc gcccagtcta
     3661 gctatcgcca tgtaagccca ctgcaagcta cctgctttct ctttgcgctt gcgttttccc
     3721 ttgtccagat agcccagtag ctgacattca tccggggtca gcaccgtttc tgcggactgg
     3781 ctttctacgt gttccgcttc ctttagcagc ccttgcgccc tgagtgcttg cggcagcgtg
     3841 aagctgtgcc tagaaatatt ttatctgatt aataagatga tcttcttgag atcgttttgg
     3901 tctgcgcgta atctcttgct ctgaaaacga aaaaaccgcc ttgcagggcg gtttttcgaa
     3961 ggttctctga gctaccaact ctttgaaccg aggtaactgg cttggaggag cgcagtcacc
     4021 aaaacttgtc ctttcagttt agccttaacc ggcgcatgac ttcaagacta actcctctaa
     4081 atcaattacc agtggctgct gccagtggtg cttttgcatg tctttccggg ttggactcaa
     4141 gacgatagtt accggataag gcgcagcggt cggactgaac ggggggttcg tgcatacagt
     4201 ccagcttgga gcgaactgcc tacccggaac tgagtgtcag gcgtggaatg agacaaacgc
     4261 ggccataaca gcggaatgac accggtaaac cgaaaggcag gaacaggaga gcgcacgagg
     4321 gagccgccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccaccactga
     4381 tttgagcgtc agatttcgtg atgcttgtca ggggggcgga gcctatggaa aaacggcttt
     4441 gccgcggccc tctcacttcc ctgttaagta tcttcctggc atcttccagg aaatctccgc
     4501 cccgttcgta agccatttcc gctcgccgca gtcgaacgac cgagcgtagc gagtcagtga
     4561 gcgaggaagc ggaatatatc ctgtatcaca tattctgctg acgcaccggt gcagcctttt
     4621 ttctcctgcc acatgaagca cttcactgac accctcatca gtgccaacat agtaagccag
     4681 tatatacact cc
//
