LOCUS       sequence-380615-        4641 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For membrane insertion study; Expresses tse5-CT (1169-1300) /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 4641)
  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 4641)
  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 (1169-1300)
FEATURES             Location/Qualifiers
     source          1..4641
                     /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(2011..2028)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2011..2027)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2020..2042)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2186..2205)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2305..2327
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2645..3439)
                     /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(2756..2775)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3366..3385
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      3835..4380
                     /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     4281..4300
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4497..4514
                     /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 gggagctcgg cccggacacc
      661 agaaatgcct gttctggaaa accgggctgc tcagggcgat attactgcac ccggcggtgc
      721 tcgccgttta acgggtgatc agactgccgc tctgcgtgat tctcttagcg ataaacctgc
      781 aaaaaatatt attttgctga ttggcgatgg gatgggggac tcggaaatta ctgccgcacg
      841 taattatgcc gaaggtgcgg gcggcttttt taaaggtata gatgccttac cgcttaccgg
      901 gcaatacact cactatgcgc tgaataaaaa aaccggcaaa ccggactacg tcaccgactc
      961 ggctgcatca gcaaccgcct ggtcaaccgg tgtcaaaacc tataacggcg cgctgggcgt
     1021 cgatattcac gaaaaagatc acccaacgat tctggaaatg gcaaaagccg caggtctggc
     1081 gaccggtaac gtttctaccg cagagttgca ggatgccacg cccgctgcgc tggtggcaca
     1141 tgtgacctcg cgcaaatgct acggtccgag cgcgaccagt gaaaaatgtc cgggtaacgc
     1201 tctggaaaaa ggcggaaaag gatcgattac cgaacagctg cttaacgctc gtgccgacgt
     1261 tacgcttggc ggcggcgcaa aaacctttgc tgaaacggca accgctggtg aatggcaggg
     1321 aaaaacgctg cgtgaacagg cacaggcgcg tggttatcag ttggtgagcg atgctgcctc
     1381 actgaattcg gtgacggaag cgaatcagca aaaacccctg cttggcctgt ttgctgacgg
     1441 caatatgcca gtgcgctggc taggaccgaa agcaacgtac catggcaata tcgataagcc
     1501 cgcagtcacc tgtacgccaa atccgcaacg taatgacagt gtaccaaccc tggcgcagat
     1561 gaccgacaaa gccattgaat tgttgagtaa aaatgagaaa ggctttttcc tgcaagttga
     1621 aggtgcgtca atcgataaac aggatcatgc tgcgaatcct tgtgggcaaa ttggcgagac
     1681 ggtcgatctc gatgaagccg tacaacgggc gctggaattc gctaaaaagg agggtaacac
     1741 gctggtcata gtcaccgctg atcacgccca cgccagccag attgttgcgc cggataccaa
     1801 agctccgggc ctcacccagg cgctaaatac caaagatggc gcagtgatgg tgatgagtta
     1861 cgggaactcc gaagaggatt cacaagaaca taccggcagt cagttgcgta ttgcggcgta
     1921 tggcccgcat gccgccaatg ttgttggact gaccgaccag accgatctct tctacaccat
     1981 gaaagccgct ctggggctga aaacggattc actggccgtc gttttacaac gtcgtgactg
     2041 ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg
     2101 gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg
     2161 cgaatggcgc taagtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     2221 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     2281 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     2341 caccgaaacg cgcgagacga aagggccgcc cttcccaaca gttgcgcagc ctgaatggcg
     2401 aatggcgctg atgtccggcg gtgcttttgc cgttacgcac caccccgtca gtagctgaac
     2461 aggagggaca gggggtgggc gaagaactcc agcatgagat ccccgcgctg gaggatcatc
     2521 cagccggcgt cccggaaaac gattccgaag cccaaccttt catagaaggc ggcggtggaa
     2581 tcgaaatctc gtgatggcag gttgggcgtc gcttggtcgg tcatttcgaa ccccagagtc
     2641 ccgctcagaa gaactcgtca agaaggcgat agaaggcgat gcgctgcgaa tcgggagcgg
     2701 cgataccgta aagcacgagg aagcggtcag cccattcgcc gccaagctct tcagcaatat
     2761 cacgggtagc caacgctatg tcctgatagc ggtccgccac acccagccgg ccacagtcga
     2821 tgaatccaga aaagcggcca ttttccacca tgatattcgg caagcaggca tcgccatggg
     2881 tcacgacgag atcctcgccg tcgggcatcc gcgccttgag cctggcgaac agttcggctg
     2941 gcgcgagccc ctgatgctct tcgtccagat catcctgatc gacaagaccg gcttccatcc
     3001 gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc gaatgggcag gtagccggat
     3061 caagcgtatg cagccgccgc attgcatcag ccatgatgga tactttctcg gcaggagcaa
     3121 ggtgagatga caggagatcc tgccccggca cttcgcccaa tagcagccag tcccttcccg
     3181 cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc cgtcgtggcc agccacgata
     3241 gccgcgctgc ctcgtcttgg agttcattca gggcaccgga caggtcggtc ttgacaaaaa
     3301 gaaccgggcg cccctgcgct gacagccgga acacggcggc atcagagcag ccgattgtct
     3361 gttgtgccca gtcatagccg aatagcctct ccacccaagc ggccggagaa cctgcgtgca
     3421 atccatcttg ttcaatcatg cgaaacgatc ctcatcctgt ctcttgatca gatcttgatc
     3481 ccctgcgcca tcagatcctt ggcggcaaga aagccatcca gtttactttg cagggcttcc
     3541 caaccttacc agagggcgcc ccagctggca attccggttc gcttgctgtc cataaaaccg
     3601 cccagtctag ctatcgccat gtaagcccac tgcaagctac ctgctttctc tttgcgcttg
     3661 cgttttccct tgtccagata gcccagtagc tgacattcat ccggggtcag caccgtttct
     3721 gcggactggc tttctacgtg ttccgcttcc tttagcagcc cttgcgccct gagtgcttgc
     3781 ggcagcgtga agctgtgcct agaaatattt tatctgatta ataagatgat cttcttgaga
     3841 tcgttttggt ctgcgcgtaa tctcttgctc tgaaaacgaa aaaaccgcct tgcagggcgg
     3901 tttttcgaag gttctctgag ctaccaactc tttgaaccga ggtaactggc ttggaggagc
     3961 gcagtcacca aaacttgtcc tttcagttta gccttaaccg gcgcatgact tcaagactaa
     4021 ctcctctaaa tcaattacca gtggctgctg ccagtggtgc ttttgcatgt ctttccgggt
     4081 tggactcaag acgatagtta ccggataagg cgcagcggtc ggactgaacg gggggttcgt
     4141 gcatacagtc cagcttggag cgaactgcct acccggaact gagtgtcagg cgtggaatga
     4201 gacaaacgcg gccataacag cggaatgaca ccggtaaacc gaaaggcagg aacaggagag
     4261 cgcacgaggg agccgccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     4321 caccactgat ttgagcgtca gatttcgtga tgcttgtcag gggggcggag cctatggaaa
     4381 aacggctttg ccgcggccct ctcacttccc tgttaagtat cttcctggca tcttccagga
     4441 aatctccgcc ccgttcgtaa gccatttccg ctcgccgcag tcgaacgacc gagcgtagcg
     4501 agtcagtgag cgaggaagcg gaatatatcc tgtatcacat attctgctga cgcaccggtg
     4561 cagccttttt tctcctgcca catgaagcac ttcactgaca ccctcatcag tgccaacata
     4621 gtaagccagt atatacactc c
//
