LOCUS       sequence-380608-        7067 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of Tse5-CT (1169-1317) in P. putida with the PelB 
            signal peptide fused at the N-terminus in biological function 
            studies.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7067)
  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 7067)
  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: pS238D1::sptse5-CT
FEATURES             Location/Qualifiers
     source          1..7067
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(19..984)
                     /codon_start=1
                     /gene="xylS"
                     /product="XylS regulator encoded by the Pseudomonas putida 
                     TOL plasmid pWWO"
                     /label=XylS
                     /note="stimulates transcription from the Pm promoter in the
                     presence of aromatic compounds such as m-toluate or 
                     benzoate"
                     /translation="MDFCLLNEKSQIFVHAEPYAVSDYVNQYVGTHSIRLPKGGRPAGR
                     LHHRIFGCLDLCRISYGGSVRVISPGLETCYHLQIILKGHCLWRGHGQEHYFAPGELLL
                     LNPDDQADLTYSEDCEKFIVKLPSVVLDRACSDNNWHKPREGIRFAARHNLQQLDGFIN
                     LLGLVCDEAEHTKSMPRVQEHYAGIIASKLLEMLGSNVSREIFSKGNPSFERVVQFIEE
                     NLKRNISLERLAELAMMSPRSLYNLFEKHAGTTPKNYIRNRKLESIRACLNDPSANVRS
                     ITEIALDYGFLHLGRFAENYRSAFGELPSDTLRQCKKEVA"
     promoter        1884..1964
                     /label=Pm promoter
                     /note="The bacterial Pm promoter is activated by XylS in 
                     the presence of benzoate or m-toluate (Marques et al., 
                     1999)."
     CDS             2018..3100
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(2037..2056)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     sig_peptide     3135..3200
                     /codon_start=1
                     /gene="pelB (fragment)"
                     /product="leader peptide for secretion"
                     /label=pelB signal sequence
                     /translation="MKYLLPTAAAGLLLLAAQPAMA"
     CDS             3666..3683
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      complement(3800..3827)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(3843..3913)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4099..4193
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(4314..4333)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4325..5140
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     oriT            5307..5415
                     /note="incP origin of transfer"
     CDS             complement(5426..6088)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      6089..6860
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     terminator      complement(6975..7061)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttaattaaac gttcgtaatc aagccacttc ctttttgcat tgacgcaggg tgtcggaagg
       61 caactcgccg aacgcgctcc tatagttttc agcgaagcgt cccaaatgta agaagccgta
      121 gtctagggct atctcagtta tactacgcac attggcactg ggatcgttca agcaggcgcg
      181 gatgctttcg agcttgcggt tgcggatgta gttcttcggc gtggtgccgg cgtgcttctc
      241 gaacaaattg tagagcgagc gtggactcat catcgccagc tccgctaacc gctcaaggct
      301 gatattccgt ttgagattct cctcaatgaa ttgaacgact cgctcgaaag acgggttacc
      361 tttgctgaaa atttcacggc tgacattgct gcccagcatt tcgagcagct tggaagcgat
      421 gatccccgca tagtgctctt ggacccgagg catcgacttt gtatgttccg cttcgtcaca
      481 aactaacccg agtagattga taaagccatc gagttgctgg agattgtgtc gcgcggcgaa
      541 acggataccc tccctcggct tgtgccaatt gttgtcactg cacgcccgat caaggaccac
      601 tgagggcaat ttaacgataa atttctcgca atcttctgaa taggtcaggt cggcttggtc
      661 atccggattg agcagcaata gttcgcccgg cgcaaaatag tgctcctggc catggccacg
      721 ccacaggcaa tggcctttga gtattatttg cagatgataa caggtctcta atccaggcga
      781 gattaccctc acgctaccgc cgtagctgat tcgacacaga tcgaggcatc cgaagattct
      841 gtggtgcagc ctgcctgccg ggcgcccgcc cttgggcagg cgaatagagt gcgtaccgac
      901 atactggtta acataatcgg agactgcata gggctcggcg tggacgaaga tctgactttt
      961 ctcgttcaat aagcaaaaat ccatagttca cggttctctt attttaatgt gggctgcttg
     1021 gtgtgatgta gaaaggcgcc aagtcgatga aaatgcatct cgacgtgatg cgtatacggg
     1081 ttacccccat tgccacgttg cgccatcctt tttgcaatca gtgaccactt ttccaagcaa
     1141 aaataacgcc aagcagaacg aagacgttct ttttaagaag cgagaacacc agaagttcgt
     1201 gctgtcgggg catggggcga cgaattggcg gataaagggg atctgctgga tattacggcc
     1261 tttttaaaga ccgtaaagaa aaataagcac aagttttatc cggcctttat tcacattctt
     1321 gcccgcctga tgaatgctca tccgtaatta cgtatggcaa tgaaagacgg tgagctggtg
     1381 atatgggata gtgttcaccc ttgttacacc gttttccatg agcaaactga aacgttttca
     1441 tcgctctgga gtgaatacca cgacgatttc cggcagtttc tacacatata ttcgcaagat
     1501 gtggcgtgtt acggtgaaaa cctggcctat ttccctaaag ggtttattga gaatatgttt
     1561 ttcgtctcag ccaatccctg ggtgagtttc accagttttg atttaaacgt ggccaatatg
     1621 gacaacttct tcgcccccgt tttcaccatg catgggaatt agcttgatct gaccaacgac
     1681 cggtagcgga gctatccaac ggcggtatac caggaaaaca cacagcaggt acatcagaac
     1741 agtaccatga ctgaagaaca aatagttttt tcctgatcca taaagcagaa cggcctgctc
     1801 catgacaaat ctggctcccc aactaatgcc ccatgcagcc agcataacca gcataaagtg
     1861 cagtgtccgg tttgataggg ataagtccag ccttgcaaga agcggataca ggagtgcaaa
     1921 aaatggctat ctctagtaag gcctacccct taggctttat gcaacagaaa caataataat
     1981 gggagtcatg accatgccta ggaggaaaaa acatatggtg aaaccagtaa cgttatacga
     2041 tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga accaggccag
     2101 ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc tgaattacat
     2161 tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg gcgttgccac
     2221 ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga
     2281 tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa
     2341 agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta actatccgct
     2401 ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg cgttatttct
     2461 tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgaag acggtacgcg
     2521 actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc
     2581 attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc tcactcgcaa
     2641 tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg gttttcaaca
     2701 aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg ccaacgatca
     2761 gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg gtgcggatat
     2821 ctcggtagtg ggatacgacg ataccgaaga cagctcatgt tatatcccgc cgttaaccac
     2881 catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc tgcaactctc
     2941 tcagggccag gcggtgaagg gcaatcagct gttgcccgtc tcactggtga aaagaaaaac
     3001 caccctggcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     3061 gctggcacga caggtttccc gactggaaag cgggcagtaa catcaccacc atcattagga
     3121 tggtggtgat gataatgaaa tacctgctgc cgaccgctgc tgctggtctg ctgctcctcg
     3181 ctgcccagcc ggcgatggcc atgggactga tcatcccgct ggtggtgatc ggcgccttcg
     3241 ccgcccgcgc cgcgatcgga gcggcgctgg gcgccggcat cgagctgggg atgcagaccg
     3301 gcaagcaggt gctggggcag atgaaggaca actgggacag cgatcgcgac ctcaccgata
     3361 tcaagtggaa gtgcatcgac atcaactgga aacacgtcgg cgcctcggcg gccatcggca
     3421 ccgtggcgcc gggcatgctc agcaccggca agaccgtggt gcagtccgcc aaggcgatca
     3481 ggaccctttc cggacaggcg gccaacaccg ccaaccgcgc ggccaagctg gccgcgcgca
     3541 aggccgccca tgccgatacc atcaagaagg ccgtggccac ccaggccgcc tggcagacgg
     3601 gcaagcagat cgtcaaatgc ccactgaagg acgaagaaga ggagtgtccg ccgcaatgac
     3661 tcgagcacca ccaccaccac cactgaggta cccggggatc ctctagagtc gacctgcagg
     3721 catgcaagct tgcggccgcg tcgtgactgg gaaaaccctg gcgcaaagcc cgccgaaagg
     3781 cgggcttttc tgttataaac gcagaaaggc ccacccgaag gtgagccagt gtgactctag
     3841 tagagagcgt tcaccgacaa aaacagataa aacgaaaggc ccagtctttc gactgagcct
     3901 ttcgttttat ttgatgcctg gctcgagctc gagaaaaaaa gcccggacga ctgttcgggc
     3961 ttgtcttttt atatgttgga aaatcagtgg caatgcaatg gcccaacaga aaatggtgaa
     4021 accagtaacg ttatactgaa tctaagtatc aatgtgagcg ctcacaagtc aacactcttt
     4081 ttgataaaac tagtcttgga ctcctgttga tagatccagt aatgacctca gaactccatc
     4141 tggatttgtt cagaacgctc ggttgccgcc gggcgttttt tattggtgag aatccagggg
     4201 tccccaataa ttacgattta aatttgtgtc tcaaaatctc tgatgttaca ttgcacaaga
     4261 taaaaatata tcatcatgaa caataaaact gtctgcttac ataaacagta atacaagggg
     4321 tgttatgagc catattcagc gtgaaacgag ctgtagccgt ccgcgtctga acagcaacat
     4381 ggatgcggat ctgtatggct ataaatgggc gcgtgataac gtgggtcaga gcggcgcgac
     4441 catttatcgt ctgtatggca aaccggatgc gccggaactg tttctgaaac atggcaaagg
     4501 cagcgtggcg aacgatgtga ccgatgaaat ggtgcgtctg aactggctga ccgaatttat
     4561 gccgctgccg accattaaac attttattcg caccccggat gatgcgtggc tgctgaccac
     4621 cgcgattccg ggcaaaaccg cgtttcaggt gctggaagaa tatccggata gcggcgaaaa
     4681 cattgtggat gcgctggccg tgtttctgcg tcgtctgcat agcattccgg tgtgcaactg
     4741 cccgtttaac agcgatcgtg tgtttcgtct ggcccaggcg cagagccgta tgaacaacgg
     4801 cctggtggat gcgagcgatt ttgatgatga acgtaacggc tggccggtgg aacaggtgtg
     4861 gaaagaaatg cataaactgc tgccgtttag cccggatagc gtggtgaccc acggcgattt
     4921 tagcctggat aacctgattt tcgatgaagg caaactgatt ggctgcattg atgtgggccg
     4981 tgtgggcatt gcggatcgtt atcaggatct ggccattctg tggaactgcc tgggcgaatt
     5041 tagcccgagc ctgcaaaaac gtctgtttca gaaatatggc attgataatc cggatatgaa
     5101 caaactgcaa tttcatctga tgctggatga atttttctaa taattaattg gaccgcggtc
     5161 cgcgcgttgt ccttttccgc tgcataaccc tgcttcgggg tcattatagc gattttttcg
     5221 gtatatccat cctttttcgc acgatataca ggattttgcc aaagggttcg tgtagacttt
     5281 ccttggtgta tccaacggcg tcagccgggc aggataggtg aagtaggccc acccgcgagc
     5341 gggtgttcct tcttcactgt cccttattcg cacctggcgg tgctcaacgg gaatcctgct
     5401 ctgcgaggct ggccgtaggc cggccctacc ggcgcggcag cgttacccgt gtcggcggct
     5461 ccaacggctc gccatcgtcc agaaaacacg gctcatcggg catcggcagg cgctgctgcc
     5521 cgcgccgttc ccattcctcc gtttcggtca aggctggcag gtctggttcc atgcccggaa
     5581 tgccgggctg gctgggcggc tcctcgccgg ggccggtcgg tagttgctgc tcgcccggat
     5641 acagggtcgg gatgcggcgc aggtcgccat gccccaacag cgattcgtcc tggtcgtcgt
     5701 gatcaaccac cacggcggca ctgaacaccg acaggcgcaa ctggtcgcgg ggctggcccc
     5761 acgccacgcg gtcattgacc acgtaggccg acacggtgcc ggggccgttg agcttcacga
     5821 cggagatcca gcgctcggcc accaagtcct tgactgcgta ttggaccgtc cgcaaagaac
     5881 gtccgatgag cttggaaagt gtcttctggc tgaccaccac ggcgttctgg tggcccatct
     5941 gcgccacgag gtgatgcagc agcattgccg ccgtgggttt cctcgcaata agcccggccc
     6001 acgcctcatg cgctttgcgt tccgtttgca cccagtgacc gggcttgttc ttggcttgaa
     6061 tgccgatttc tctggactgc gtggccatgc ttatctccat gcggtagggg tgccgcacgg
     6121 ttgcggcacc atgcgcaatc agctgcaact tttcggcagc gcgacaacaa ttatgcgttg
     6181 cgtaaaagtg gcagtcaatt acagattttc tttaacctac gcaatgagct attgcggggg
     6241 gtgccgcaat gagctgttgc gtacccccct tttttaagtt gttgattttt aagtctttcg
     6301 catttcgccc tatatctagt tctttggtgc ccaaagaagg gcacccctgc ggggttcccc
     6361 cacgccttcg gcgcggctcc ccctccggca aaaagtggcc cctccggggc ttgttgatcg
     6421 actgcgcggc cttcggcctt gcccaaggtg gcgctgcccc cttggaaccc ccgcactcgc
     6481 cgccgtgagg ctcggggggc aggcgggcgg gcttcgccct tcgactgccc ccactcgcat
     6541 aggcttgggt cgttccaggc gcgtcaaggc caagccgctg cgcggtcgct gcgcgagcct
     6601 tgacccgcct tccacttggt gtccaaccgg caagcgaagc gcgcaggccg caggccggag
     6661 gcttttcccc agagaaaatt aaaaaaattg atggggcaag gccgcaggcc gcgcagttgg
     6721 agccggtggg tatgtggtcg aaggctgggt agccggtggg caatccctgt ggtcaagctc
     6781 gtgggcaggc gcagcctgtc catcagcttg tccagcaggg ttgtccacgg gccgagcgaa
     6841 gcgagccagc cggtggccgc tcgcggccat cgtccacata tccacgggct ggcaagggag
     6901 cgcagcgacc gcgcagggcg aagcccggag agcaagcccg tagggggggc gcgcccagct
     6961 gtctagggcg gcggatttgt cctactcagg agagcgttca ccgacaaaca acagataaaa
     7021 cgaaaggccc agtctttcga ctgagccttt cgttttattt gatgcct
//
