LOCUS       sequence-380612-        4548 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For membrane insertion study; Expresses tse5-CT (1169-1269) /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 4548)
  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 4548)
  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 (1169-1269)
FEATURES             Location/Qualifiers
     source          1..4548
                     /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(1918..1935)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1918..1934)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1927..1949)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2093..2112)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2212..2234
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2552..3346)
                     /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(2663..2682)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3273..3292
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      3742..4287
                     /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     4188..4207
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4404..4421
                     /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 caccgcgggg agctcggccc ggacaccaga aatgcctgtt ctggaaaacc gggctgctca
      601 gggcgatatt actgcacccg gcggtgctcg ccgtttaacg ggtgatcaga ctgccgctct
      661 gcgtgattct cttagcgata aacctgcaaa aaatattatt ttgctgattg gcgatgggat
      721 gggggactcg gaaattactg ccgcacgtaa ttatgccgaa ggtgcgggcg gcttttttaa
      781 aggtatagat gccttaccgc ttaccgggca atacactcac tatgcgctga ataaaaaaac
      841 cggcaaaccg gactacgtca ccgactcggc tgcatcagca accgcctggt caaccggtgt
      901 caaaacctat aacggcgcgc tgggcgtcga tattcacgaa aaagatcacc caacgattct
      961 ggaaatggca aaagccgcag gtctggcgac cggtaacgtt tctaccgcag agttgcagga
     1021 tgccacgccc gctgcgctgg tggcacatgt gacctcgcgc aaatgctacg gtccgagcgc
     1081 gaccagtgaa aaatgtccgg gtaacgctct ggaaaaaggc ggaaaaggat cgattaccga
     1141 acagctgctt aacgctcgtg ccgacgttac gcttggcggc ggcgcaaaaa cctttgctga
     1201 aacggcaacc gctggtgaat ggcagggaaa aacgctgcgt gaacaggcac aggcgcgtgg
     1261 ttatcagttg gtgagcgatg ctgcctcact gaattcggtg acggaagcga atcagcaaaa
     1321 acccctgctt ggcctgtttg ctgacggcaa tatgccagtg cgctggctag gaccgaaagc
     1381 aacgtaccat ggcaatatcg ataagcccgc agtcacctgt acgccaaatc cgcaacgtaa
     1441 tgacagtgta ccaaccctgg cgcagatgac cgacaaagcc attgaattgt tgagtaaaaa
     1501 tgagaaaggc tttttcctgc aagttgaagg tgcgtcaatc gataaacagg atcatgctgc
     1561 gaatccttgt gggcaaattg gcgagacggt cgatctcgat gaagccgtac aacgggcgct
     1621 ggaattcgct aaaaaggagg gtaacacgct ggtcatagtc accgctgatc acgcccacgc
     1681 cagccagatt gttgcgccgg ataccaaagc tccgggcctc acccaggcgc taaataccaa
     1741 agatggcgca gtgatggtga tgagttacgg gaactccgaa gaggattcac aagaacatac
     1801 cggcagtcag ttgcgtattg cggcgtatgg cccgcatgcc gccaatgttg ttggactgac
     1861 cgaccagacc gatctcttct acaccatgaa agccgctctg gggctgaaaa cggattcact
     1921 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct
     1981 tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc
     2041 ttcccaacag ttgcgcagcc tgaatggcga atggcgctaa gtgcactctc agtacaatct
     2101 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     2161 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     2221 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggccgccctt
     2281 cccaacagtt gcgcagcctg aatggcgaat ggcgctgatg tccggcggtg cttttgccgt
     2341 tacgcaccac cccgtcagta gctgaacagg agggacaggg ggtgggcgaa gaactccagc
     2401 atgagatccc cgcgctggag gatcatccag ccggcgtccc ggaaaacgat tccgaagccc
     2461 aacctttcat agaaggcggc ggtggaatcg aaatctcgtg atggcaggtt gggcgtcgct
     2521 tggtcggtca tttcgaaccc cagagtcccg ctcagaagaa ctcgtcaaga aggcgataga
     2581 aggcgatgcg ctgcgaatcg ggagcggcga taccgtaaag cacgaggaag cggtcagccc
     2641 attcgccgcc aagctcttca gcaatatcac gggtagccaa cgctatgtcc tgatagcggt
     2701 ccgccacacc cagccggcca cagtcgatga atccagaaaa gcggccattt tccaccatga
     2761 tattcggcaa gcaggcatcg ccatgggtca cgacgagatc ctcgccgtcg ggcatccgcg
     2821 ccttgagcct ggcgaacagt tcggctggcg cgagcccctg atgctcttcg tccagatcat
     2881 cctgatcgac aagaccggct tccatccgag tacgtgctcg ctcgatgcga tgtttcgctt
     2941 ggtggtcgaa tgggcaggta gccggatcaa gcgtatgcag ccgccgcatt gcatcagcca
     3001 tgatggatac tttctcggca ggagcaaggt gagatgacag gagatcctgc cccggcactt
     3061 cgcccaatag cagccagtcc cttcccgctt cagtgacaac gtcgagcaca gctgcgcaag
     3121 gaacgcccgt cgtggccagc cacgatagcc gcgctgcctc gtcttggagt tcattcaggg
     3181 caccggacag gtcggtcttg acaaaaagaa ccgggcgccc ctgcgctgac agccggaaca
     3241 cggcggcatc agagcagccg attgtctgtt gtgcccagtc atagccgaat agcctctcca
     3301 cccaagcggc cggagaacct gcgtgcaatc catcttgttc aatcatgcga aacgatcctc
     3361 atcctgtctc ttgatcagat cttgatcccc tgcgccatca gatccttggc ggcaagaaag
     3421 ccatccagtt tactttgcag ggcttcccaa ccttaccaga gggcgcccca gctggcaatt
     3481 ccggttcgct tgctgtccat aaaaccgccc agtctagcta tcgccatgta agcccactgc
     3541 aagctacctg ctttctcttt gcgcttgcgt tttcccttgt ccagatagcc cagtagctga
     3601 cattcatccg gggtcagcac cgtttctgcg gactggcttt ctacgtgttc cgcttccttt
     3661 agcagccctt gcgccctgag tgcttgcggc agcgtgaagc tgtgcctaga aatattttat
     3721 ctgattaata agatgatctt cttgagatcg ttttggtctg cgcgtaatct cttgctctga
     3781 aaacgaaaaa accgccttgc agggcggttt ttcgaaggtt ctctgagcta ccaactcttt
     3841 gaaccgaggt aactggcttg gaggagcgca gtcaccaaaa cttgtccttt cagtttagcc
     3901 ttaaccggcg catgacttca agactaactc ctctaaatca attaccagtg gctgctgcca
     3961 gtggtgcttt tgcatgtctt tccgggttgg actcaagacg atagttaccg gataaggcgc
     4021 agcggtcgga ctgaacgggg ggttcgtgca tacagtccag cttggagcga actgcctacc
     4081 cggaactgag tgtcaggcgt ggaatgagac aaacgcggcc ataacagcgg aatgacaccg
     4141 gtaaaccgaa aggcaggaac aggagagcgc acgagggagc cgccaggggg aaacgcctgg
     4201 tatctttata gtcctgtcgg gtttcgccac cactgatttg agcgtcagat ttcgtgatgc
     4261 ttgtcagggg ggcggagcct atggaaaaac ggctttgccg cggccctctc acttccctgt
     4321 taagtatctt cctggcatct tccaggaaat ctccgccccg ttcgtaagcc atttccgctc
     4381 gccgcagtcg aacgaccgag cgtagcgagt cagtgagcga ggaagcggaa tatatcctgt
     4441 atcacatatt ctgctgacgc accggtgcag ccttttttct cctgccacat gaagcacttc
     4501 actgacaccc tcatcagtgc caacatagta agccagtata tacactcc
//
