LOCUS       sequence-380606-        4584 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For membrane insertion study; Expresses tse5-CT (1169-1281) /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 4584)
  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 4584)
  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 (1169-1281)
FEATURES             Location/Qualifiers
     source          1..4584
                     /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(1954..1971)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1954..1970)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1963..1985)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2129..2148)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2248..2270
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2588..3382)
                     /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(2699..2718)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3309..3328
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      3778..4323
                     /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     4224..4243
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4440..4457
                     /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 gcggggagct cggcccggac
      601 accagaaatg cctgttctgg aaaaccgggc tgctcagggc gatattactg cacccggcgg
      661 tgctcgccgt ttaacgggtg atcagactgc cgctctgcgt gattctctta gcgataaacc
      721 tgcaaaaaat attattttgc tgattggcga tgggatgggg gactcggaaa ttactgccgc
      781 acgtaattat gccgaaggtg cgggcggctt ttttaaaggt atagatgcct taccgcttac
      841 cgggcaatac actcactatg cgctgaataa aaaaaccggc aaaccggact acgtcaccga
      901 ctcggctgca tcagcaaccg cctggtcaac cggtgtcaaa acctataacg gcgcgctggg
      961 cgtcgatatt cacgaaaaag atcacccaac gattctggaa atggcaaaag ccgcaggtct
     1021 ggcgaccggt aacgtttcta ccgcagagtt gcaggatgcc acgcccgctg cgctggtggc
     1081 acatgtgacc tcgcgcaaat gctacggtcc gagcgcgacc agtgaaaaat gtccgggtaa
     1141 cgctctggaa aaaggcggaa aaggatcgat taccgaacag ctgcttaacg ctcgtgccga
     1201 cgttacgctt ggcggcggcg caaaaacctt tgctgaaacg gcaaccgctg gtgaatggca
     1261 gggaaaaacg ctgcgtgaac aggcacaggc gcgtggttat cagttggtga gcgatgctgc
     1321 ctcactgaat tcggtgacgg aagcgaatca gcaaaaaccc ctgcttggcc tgtttgctga
     1381 cggcaatatg ccagtgcgct ggctaggacc gaaagcaacg taccatggca atatcgataa
     1441 gcccgcagtc acctgtacgc caaatccgca acgtaatgac agtgtaccaa ccctggcgca
     1501 gatgaccgac aaagccattg aattgttgag taaaaatgag aaaggctttt tcctgcaagt
     1561 tgaaggtgcg tcaatcgata aacaggatca tgctgcgaat ccttgtgggc aaattggcga
     1621 gacggtcgat ctcgatgaag ccgtacaacg ggcgctggaa ttcgctaaaa aggagggtaa
     1681 cacgctggtc atagtcaccg ctgatcacgc ccacgccagc cagattgttg cgccggatac
     1741 caaagctccg ggcctcaccc aggcgctaaa taccaaagat ggcgcagtga tggtgatgag
     1801 ttacgggaac tccgaagagg attcacaaga acataccggc agtcagttgc gtattgcggc
     1861 gtatggcccg catgccgcca atgttgttgg actgaccgac cagaccgatc tcttctacac
     1921 catgaaagcc gctctggggc tgaaaacgga ttcactggcc gtcgttttac aacgtcgtga
     1981 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     2041 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     2101 tggcgaatgg cgctaagtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     2161 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     2221 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     2281 catcaccgaa acgcgcgaga cgaaagggcc gcccttccca acagttgcgc agcctgaatg
     2341 gcgaatggcg ctgatgtccg gcggtgcttt tgccgttacg caccaccccg tcagtagctg
     2401 aacaggaggg acagggggtg ggcgaagaac tccagcatga gatccccgcg ctggaggatc
     2461 atccagccgg cgtcccggaa aacgattccg aagcccaacc tttcatagaa ggcggcggtg
     2521 gaatcgaaat ctcgtgatgg caggttgggc gtcgcttggt cggtcatttc gaaccccaga
     2581 gtcccgctca gaagaactcg tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag
     2641 cggcgatacc gtaaagcacg aggaagcggt cagcccattc gccgccaagc tcttcagcaa
     2701 tatcacgggt agccaacgct atgtcctgat agcggtccgc cacacccagc cggccacagt
     2761 cgatgaatcc agaaaagcgg ccattttcca ccatgatatt cggcaagcag gcatcgccat
     2821 gggtcacgac gagatcctcg ccgtcgggca tccgcgcctt gagcctggcg aacagttcgg
     2881 ctggcgcgag cccctgatgc tcttcgtcca gatcatcctg atcgacaaga ccggcttcca
     2941 tccgagtacg tgctcgctcg atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg
     3001 gatcaagcgt atgcagccgc cgcattgcat cagccatgat ggatactttc tcggcaggag
     3061 caaggtgaga tgacaggaga tcctgccccg gcacttcgcc caatagcagc cagtcccttc
     3121 ccgcttcagt gacaacgtcg agcacagctg cgcaaggaac gcccgtcgtg gccagccacg
     3181 atagccgcgc tgcctcgtct tggagttcat tcagggcacc ggacaggtcg gtcttgacaa
     3241 aaagaaccgg gcgcccctgc gctgacagcc ggaacacggc ggcatcagag cagccgattg
     3301 tctgttgtgc ccagtcatag ccgaatagcc tctccaccca agcggccgga gaacctgcgt
     3361 gcaatccatc ttgttcaatc atgcgaaacg atcctcatcc tgtctcttga tcagatcttg
     3421 atcccctgcg ccatcagatc cttggcggca agaaagccat ccagtttact ttgcagggct
     3481 tcccaacctt accagagggc gccccagctg gcaattccgg ttcgcttgct gtccataaaa
     3541 ccgcccagtc tagctatcgc catgtaagcc cactgcaagc tacctgcttt ctctttgcgc
     3601 ttgcgttttc ccttgtccag atagcccagt agctgacatt catccggggt cagcaccgtt
     3661 tctgcggact ggctttctac gtgttccgct tcctttagca gcccttgcgc cctgagtgct
     3721 tgcggcagcg tgaagctgtg cctagaaata ttttatctga ttaataagat gatcttcttg
     3781 agatcgtttt ggtctgcgcg taatctcttg ctctgaaaac gaaaaaaccg ccttgcaggg
     3841 cggtttttcg aaggttctct gagctaccaa ctctttgaac cgaggtaact ggcttggagg
     3901 agcgcagtca ccaaaacttg tcctttcagt ttagccttaa ccggcgcatg acttcaagac
     3961 taactcctct aaatcaatta ccagtggctg ctgccagtgg tgcttttgca tgtctttccg
     4021 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcggactga acggggggtt
     4081 cgtgcataca gtccagcttg gagcgaactg cctacccgga actgagtgtc aggcgtggaa
     4141 tgagacaaac gcggccataa cagcggaatg acaccggtaa accgaaaggc aggaacagga
     4201 gagcgcacga gggagccgcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     4261 cgccaccact gatttgagcg tcagatttcg tgatgcttgt caggggggcg gagcctatgg
     4321 aaaaacggct ttgccgcggc cctctcactt ccctgttaag tatcttcctg gcatcttcca
     4381 ggaaatctcc gccccgttcg taagccattt ccgctcgccg cagtcgaacg accgagcgta
     4441 gcgagtcagt gagcgaggaa gcggaatata tcctgtatca catattctgc tgacgcaccg
     4501 gtgcagcctt ttttctcctg ccacatgaag cacttcactg acaccctcat cagtgccaac
     4561 atagtaagcc agtatataca ctcc
//
