LOCUS       sequence-380607-        4656 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For membrane insertion study; Expresses sptse5-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 4656)
  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 4656)
  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 (1169-1281)
FEATURES             Location/Qualifiers
     source          1..4656
                     /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(1807..1824)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1807..1823)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1816..1838)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1982..2001)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2101..2123
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2441..3235)
                     /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(2552..2571)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3162..3181
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      3631..4176
                     /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     4077..4096
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4293..4310
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4547..4568
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4583..4588,4589..4606,4607..4613)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4621..4637
                     /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     4626..4648
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     join(4645..4656,1..5)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(4645..4656,1..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 catgttggtg cgagcgcggc
      301 gattggtacc gttgcgccgg gcatgctgag caccggtaaa accgtggttc agagcgcgaa
      361 ggcgattcgt accctgagcg gtcaggcggc gaacaccgcg aaccgtgcgg cgaaactggc
      421 ggcgcgtaag gcggcgggga gctcggcccg gacaccagaa atgcctgttc tggaaaaccg
      481 ggctgctcag ggcgatatta ctgcacccgg cggtgctcgc cgtttaacgg gtgatcagac
      541 tgccgctctg cgtgattctc ttagcgataa acctgcaaaa aatattattt tgctgattgg
      601 cgatgggatg ggggactcgg aaattactgc cgcacgtaat tatgccgaag gtgcgggcgg
      661 cttttttaaa ggtatagatg ccttaccgct taccgggcaa tacactcact atgcgctgaa
      721 taaaaaaacc ggcaaaccgg actacgtcac cgactcggct gcatcagcaa ccgcctggtc
      781 aaccggtgtc aaaacctata acggcgcgct gggcgtcgat attcacgaaa aagatcaccc
      841 aacgattctg gaaatggcaa aagccgcagg tctggcgacc ggtaacgttt ctaccgcaga
      901 gttgcaggat gccacgcccg ctgcgctggt ggcacatgtg acctcgcgca aatgctacgg
      961 tccgagcgcg accagtgaaa aatgtccggg taacgctctg gaaaaaggcg gaaaaggatc
     1021 gattaccgaa cagctgctta acgctcgtgc cgacgttacg cttggcggcg gcgcaaaaac
     1081 ctttgctgaa acggcaaccg ctggtgaatg gcagggaaaa acgctgcgtg aacaggcaca
     1141 ggcgcgtggt tatcagttgg tgagcgatgc tgcctcactg aattcggtga cggaagcgaa
     1201 tcagcaaaaa cccctgcttg gcctgtttgc tgacggcaat atgccagtgc gctggctagg
     1261 accgaaagca acgtaccatg gcaatatcga taagcccgca gtcacctgta cgccaaatcc
     1321 gcaacgtaat gacagtgtac caaccctggc gcagatgacc gacaaagcca ttgaattgtt
     1381 gagtaaaaat gagaaaggct ttttcctgca agttgaaggt gcgtcaatcg ataaacagga
     1441 tcatgctgcg aatccttgtg ggcaaattgg cgagacggtc gatctcgatg aagccgtaca
     1501 acgggcgctg gaattcgcta aaaaggaggg taacacgctg gtcatagtca ccgctgatca
     1561 cgcccacgcc agccagattg ttgcgccgga taccaaagct ccgggcctca cccaggcgct
     1621 aaataccaaa gatggcgcag tgatggtgat gagttacggg aactccgaag aggattcaca
     1681 agaacatacc ggcagtcagt tgcgtattgc ggcgtatggc ccgcatgccg ccaatgttgt
     1741 tggactgacc gaccagaccg atctcttcta caccatgaaa gccgctctgg ggctgaaaac
     1801 ggattcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact
     1861 taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac
     1921 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgctaag tgcactctca
     1981 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     2041 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     2101 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg
     2161 gccgcccttc ccaacagttg cgcagcctga atggcgaatg gcgctgatgt ccggcggtgc
     2221 ttttgccgtt acgcaccacc ccgtcagtag ctgaacagga gggacagggg gtgggcgaag
     2281 aactccagca tgagatcccc gcgctggagg atcatccagc cggcgtcccg gaaaacgatt
     2341 ccgaagccca acctttcata gaaggcggcg gtggaatcga aatctcgtga tggcaggttg
     2401 ggcgtcgctt ggtcggtcat ttcgaacccc agagtcccgc tcagaagaac tcgtcaagaa
     2461 ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat accgtaaagc acgaggaagc
     2521 ggtcagccca ttcgccgcca agctcttcag caatatcacg ggtagccaac gctatgtcct
     2581 gatagcggtc cgccacaccc agccggccac agtcgatgaa tccagaaaag cggccatttt
     2641 ccaccatgat attcggcaag caggcatcgc catgggtcac gacgagatcc tcgccgtcgg
     2701 gcatccgcgc cttgagcctg gcgaacagtt cggctggcgc gagcccctga tgctcttcgt
     2761 ccagatcatc ctgatcgaca agaccggctt ccatccgagt acgtgctcgc tcgatgcgat
     2821 gtttcgcttg gtggtcgaat gggcaggtag ccggatcaag cgtatgcagc cgccgcattg
     2881 catcagccat gatggatact ttctcggcag gagcaaggtg agatgacagg agatcctgcc
     2941 ccggcacttc gcccaatagc agccagtccc ttcccgcttc agtgacaacg tcgagcacag
     3001 ctgcgcaagg aacgcccgtc gtggccagcc acgatagccg cgctgcctcg tcttggagtt
     3061 cattcagggc accggacagg tcggtcttga caaaaagaac cgggcgcccc tgcgctgaca
     3121 gccggaacac ggcggcatca gagcagccga ttgtctgttg tgcccagtca tagccgaata
     3181 gcctctccac ccaagcggcc ggagaacctg cgtgcaatcc atcttgttca atcatgcgaa
     3241 acgatcctca tcctgtctct tgatcagatc ttgatcccct gcgccatcag atccttggcg
     3301 gcaagaaagc catccagttt actttgcagg gcttcccaac cttaccagag ggcgccccag
     3361 ctggcaattc cggttcgctt gctgtccata aaaccgccca gtctagctat cgccatgtaa
     3421 gcccactgca agctacctgc tttctctttg cgcttgcgtt ttcccttgtc cagatagccc
     3481 agtagctgac attcatccgg ggtcagcacc gtttctgcgg actggctttc tacgtgttcc
     3541 gcttccttta gcagcccttg cgccctgagt gcttgcggca gcgtgaagct gtgcctagaa
     3601 atattttatc tgattaataa gatgatcttc ttgagatcgt tttggtctgc gcgtaatctc
     3661 ttgctctgaa aacgaaaaaa ccgccttgca gggcggtttt tcgaaggttc tctgagctac
     3721 caactctttg aaccgaggta actggcttgg aggagcgcag tcaccaaaac ttgtcctttc
     3781 agtttagcct taaccggcgc atgacttcaa gactaactcc tctaaatcaa ttaccagtgg
     3841 ctgctgccag tggtgctttt gcatgtcttt ccgggttgga ctcaagacga tagttaccgg
     3901 ataaggcgca gcggtcggac tgaacggggg gttcgtgcat acagtccagc ttggagcgaa
     3961 ctgcctaccc ggaactgagt gtcaggcgtg gaatgagaca aacgcggcca taacagcgga
     4021 atgacaccgg taaaccgaaa ggcaggaaca ggagagcgca cgagggagcc gccaggggga
     4081 aacgcctggt atctttatag tcctgtcggg tttcgccacc actgatttga gcgtcagatt
     4141 tcgtgatgct tgtcaggggg gcggagccta tggaaaaacg gctttgccgc ggccctctca
     4201 cttccctgtt aagtatcttc ctggcatctt ccaggaaatc tccgccccgt tcgtaagcca
     4261 tttccgctcg ccgcagtcga acgaccgagc gtagcgagtc agtgagcgag gaagcggaat
     4321 atatcctgta tcacatattc tgctgacgca ccggtgcagc cttttttctc ctgccacatg
     4381 aagcacttca ctgacaccct catcagtgcc aacatagtaa gccagtatat acactccgct
     4441 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     4501 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
     4561 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
     4621 ttgtgagcgg ataacaattt cacacaggaa acagct
//
