LOCUS       sequence-273744-        7592 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Enables high-efficiency recombineering in P. aeruginosa.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7592)
  AUTHORS   Wannier TM, Nyerges A, Kuchwara HM, Czikkely M, Balogh D, Filsinger 
            GT, Borders NC, Gregg CJ, Lajoie MJ, Rios X, Pal C, Church GM
  TITLE     Improved bacterial recombineering by parallelized protein discovery.
  JOURNAL   Proc Natl Acad Sci U S A. 2020 Jun 16;117(24):13689-13698. doi: 
            10.1073/pnas.2001588117. Epub 2020 May 28.
  PUBMED    32467157
REFERENCE   2  (bases 1 to 7592)
  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"; doi: 
            "10.1073/pnas.2001588117"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2020-06-16- 16"; volume: "117"; issue: "24"; pages: 
            "13689-13698"
COMMENT     Sequence Label: pORTMAGE-Pa1
FEATURES             Location/Qualifiers
     source          1..7592
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        224..304
                     /label=Pm promoter
                     /note="The bacterial Pm promoter is activated by XylS in 
                     the presence of benzoate or m-toluate (Marques et al., 
                     1999)."
     RBS             320..328
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     terminator      3146..3180
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     promoter        join(3221..3226,3227..3243,3244..3249)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             3438..3971
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     oriT            4182..4290
                     /note="incP origin of transfer"
     CDS             complement(4301..4963)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGETLEPPTRVTLPRR"
     rep_origin      4964..5735
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             complement(5951..6916)
                     /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"
ORIGIN
        1 agcttgatct gaccaacgac cggtagcgga gctatccaac ggcggtatac caggaaaaca
       61 cacagcaggt acatcagaac agtaccatga ctgaagaaca aatagttttt tcctgatcca
      121 taaagcagaa cggcctgctc catgacaaat ctggctcccc aactaatgcc ccatgcagcc
      181 agcataacca gcataaagtg cagtgtccgg tttgataggg ataagtccag ccttgcaaga
      241 agcggataca ggagtgcaaa aaatggctat ctctagtaag gcctacccct taggctttat
      301 gcaatgatag ccgccaattt aaggaggttt tttcatggga actgcactaa caccgctcct
      361 gacgaagttc gccacgcgct acgagatggg taccacgcct gaagaagtgg cgaacacgct
      421 caagcagacc tgtttcaagg gccaggtcaa tgattcgcag atggtcgccc tgctgatcgt
      481 ggcagaccag tacaagctta accccttcac caaggagttg tacgcattcc ccgacaagaa
      541 caacggcatc gtgccggttg ttggtgtgga tggctgggct cggatcatca acgagaaccc
      601 acagttcgat ggcatggaat tctcaatgga ccagcaggga accgaatgca cctgcaagat
      661 ctatcggaag gaccgcagcc atgccatcag cgcgactgag tacatggccg agtgcaagcg
      721 gaacacccag ccttggcagt cccatccgcg ccggatgctt cgccacaagg caatgatcca
      781 gtgcgcacgc ctcgcgttcg ggttcgccgg catatacgac caggacgagg ccgagcggat
      841 cgttgaacga gacgtcactc ccgcagagca gtacgaggac gtcagcgagg cgatctgcct
      901 gatcaaggac tccccaacaa tggaagactt gcaggcagca ttcagcaatg cctggaaagc
      961 ctataagaca aagggcgcgc gcgaccagct tacggcggcc aaggaccagc ggaagaaaga
     1021 attgctggac gcccctatag acgttgaatt cgaggagacc ggcgatgatc gagcagcgta
     1081 acaaaaaata ggaggaaaaa catatgagtg aagcaccgcg tatccagctg ctgagtccgc
     1141 ggctggcgaa ccagatcgcc gccggtgagg tggtcgagcg tcccgcctcg gtggccaagg
     1201 aactcctgaa aaacagcctt gacgccggtt cccggcgcat tgatgtggag gtcgagcagg
     1261 gcggcatcaa gttgctgcga gtgcgcgacg acggtcgcgg catccccgcc gacgacctgc
     1321 cgctggccct ggctcgccac gccaccagca agatccgcga gctggaagac ctggagcggg
     1381 tgatgagcct cggattccgt ggcgaggcgc tggcctcgat cagctcggta gcgcgcctga
     1441 ccatgacctc gcgtaccgcc gacgccggcg aagcctggca ggtggaaacc gagggccgcg
     1501 acatgcagcc gcgggtacag ccggcggcgc acccggtggg gaccagcgtc gaggttcgcg
     1561 acctgttctt caacaccccg gcgcggcgca agttcctgcg tgccgagaag accgagttcg
     1621 accatctgca ggaggtcatc aagcgcctgg cgctggcccg cttcgacgtg gctttccacc
     1681 tgcgccacaa cggcaagacc atcttcgccc tgcacgaggc gcgagacgag ctggcccgcg
     1741 cgcgccgggt cggcgcggtg tgcggccagg cattcctcga gcaggcgctg ccgatcgagg
     1801 tcgagcgcaa cggcctgcac ctgtggggct gggtcggctt gccgaccttc tcccgcagcc
     1861 agccggacct gcagtacttc tatgtgaacg ggcgcatggt gcgcgacaag ctggtcgccc
     1921 acgcggtgcg ccaggcttat cgcgacgtgc tgtacaacgg ccggcatccg accttcgtgc
     1981 tgttcttcga agtcgatccg gcggtggtgg acgtcaacgt gcacccgacc aagcacgaag
     2041 ttcgcttccg tgacagccgg atggtccatg acttcctcta tggcaccctg catcgtgcgc
     2101 tgggcgaggt ccgtcccgac gaccagctcg cgccgcccgg ggcgaccagc ctgaccgagc
     2161 cgcgtccgac cggcgctgcc gccggcgagt tcggcccgca gggcgagatg cgcctggccg
     2221 agagcgtgct ggagagtccg gcggcacggg tgggctggag tggcggctca tcggcctccg
     2281 gcggctcctc cggctattcc gcctataccc ggccggaggc gccgccctct ctggccgagg
     2341 ccggtggcgc ctacaaggcc tacttcgcgc cgttacccgc gggcgaggcg ccagccgcgt
     2401 tgcccgagag tgcccaggac atcccgccgc tgggttacgc cctggcgcag ctcaagggca
     2461 tctacattct cgccgagaac gcccacggcc tggtgctggt ggacatgcac gcggcccacg
     2521 agcggatcac ctacgagcgc ctcaaggtgg cgatggccag cgaaggcttg cgtgggcagc
     2581 cgttgctggt gccggagtcg atcgcggtca gcgagcgcga ggcggattgc gcggaagagc
     2641 attccagctg gttccagcgc ctgggcttcg agctgcagcg cctggggccg gaaagcctgg
     2701 cgatccggca gatccctgcg ctgctcaagc aggcggaggc gacccagttg gtgcgcgatg
     2761 tcatcgccga cctgctggaa tacggcacca gtgaccgtat ccaggcgcac ctgaacgagc
     2821 tgctcggcac catggcctgc catggcgcgg tgcgcgccaa ccggcgcctg accttgccgg
     2881 aaatgaacgc cctgttgcgc gacatggaaa tcaccgaacg cagcggccag tgcaaccatg
     2941 ggcgtccgac ctggacccag ttgggcctgg acgagctgga caagctcttc ctgcgcggac
     3001 gctgatctag agtcgacctg caggcatgca agcttgcggc cgcgtcgtga ctgggaaaac
     3061 cctggcgact agtcttggac tcctgttgat agatccagta atgacctcag aactccatct
     3121 ggatttgttc agacatctgg atttgttcag aacgctcggt tgccgccggg cgttttttat
     3181 tggtgagaat ccaggggtcc ccaataatta cgatttaaat ttgacataag cctgttcggt
     3241 tcgtaaactg taatgcaagt agcgtatgcg ctcacgcaac tggtccagaa ccttgaccga
     3301 acgcagcggt ggtaacggcg cagtggcggt tttcatggct tgttatgact gtttttttgt
     3361 acagcctatg cctcgggcat ccaagcagca agcgcgttac gccgtgggtc gatgtttgat
     3421 gttatggagc agcaacgatg ttacgcagca gcaacgatgt tacgcagcag ggcagtcgcc
     3481 ctaaaacaaa gttaggtggc tcaagtatgg gcatcattcg cacatgtagg ctcggccctg
     3541 accaagtcaa atccatgcgg gctgctcttg atcttttcgg tcgtgagttc ggagacgtag
     3601 ccacctactc ccaacatcag ccggactccg attacctcgg gaacttgctc cgtagtaaga
     3661 cattcatcgc gcttgctgcc ttcgaccaag aagcggttgt tggcgctctc gcggcttacg
     3721 ttctgcccaa gtttgagcag ccgcgtagtg agatctatat ctatgatctc gcagtctccg
     3781 gagagcaccg gaggcagggc attgccaccg cgctcatcaa tctcctcaag catgaggcca
     3841 acgcgcttgg tgcttatgtg atctacgtgc aagcagatta cggtgacgat cccgcagtgg
     3901 ctctctatac aaagttgggc atacgggaag aagtgatgca ctttgatatc gacccaagta
     3961 ccgccaccta acaattcgtt caagccgaga tcggcttccc ggccgcggag ttgttcggta
     4021 aattggacaa cggtccgcgc gttgtccttt tccgctgcat aaccctgctt cggggtcatt
     4081 atagcgattt tttcggtata tccatccttt ttcgcacgat atacaggatt ttgccaaagg
     4141 gttcgtgtag actttccttg gtgtatccaa cggcgtcagc cgggcaggat aggtgaagta
     4201 ggcccacccg cgagcgggtg ttccttcttc actgtccctt attcgcacct ggcggtgctc
     4261 aacgggaatc ctgctctgcg aggctggccg taggccggcc ctaccggcgc ggcagcgtta
     4321 cccgtgtcgg cggctccaac gtctcgccat cgtccagaaa acacggctca tcgggcatcg
     4381 gcaggcgctg ctgcccgcgc cgttcccatt cctccgtttc ggtcaaggct ggcaggtctg
     4441 gttccatgcc cggaatgccg ggctggctgg gcggctcctc gccggggccg gtcggtagtt
     4501 gctgctcgcc cggatacagg gtcgggatgc ggcgcaggtc gccatgcccc aacagcgatt
     4561 cgtcctggtc gtcgtgatca accaccacgg cggcactgaa caccgacagg cgcaactggt
     4621 cgcggggctg gccccacgcc acgcggtcat tgaccacgta ggccgacacg gtgccggggc
     4681 cgttgagctt cacgacggag atccagcgct cggccaccaa gtccttgact gcgtattgga
     4741 ccgtccgcaa agaacgtccg atgagcttgg aaagtgtctt ctggctgacc accacggcgt
     4801 tctggtggcc catctgcgcc acgaggtgat gcagcagcat tgccgccgtg ggtttcctcg
     4861 caataagccc ggcccacgcc tcatgcgctt tgcgttccgt ttgcacccag tgaccgggct
     4921 tgttcttggc ttgaatgccg atttctctgg actgcgtggc catgcttatc tccatgcggt
     4981 aggggtgccg cacggttgcg gcaccatgcg caatcagctg caacttttcg gcagcgcgac
     5041 aacaattatg cgttgcgtaa aagtggcagt caattacaga ttttctttaa cctacgcaat
     5101 gagctattgc ggggggtgcc gcaatgagct gttgcgtacc cccctttttt aagttgttga
     5161 tttttaagtc tttcgcattt cgccctatat ctagttcttt ggtgcccaaa gaagggcacc
     5221 cctgcggggt tcccccacgc cttcggcgcg gctccccctc cggcaaaaag tggcccctcc
     5281 ggggcttgtt gatcgactgc gcggccttcg gccttgccca aggtggcgct gcccccttgg
     5341 aacccccgca ctcgccgccg tgaggctcgg ggggcaggcg ggcgggcttc gcccttcgac
     5401 tgcccccact cgcataggct tgggtcgttc caggcgcgtc aaggccaagc cgctgcgcgg
     5461 tcgctgcgcg agccttgacc cgccttccac ttggtgtcca accggcaagc gaagcgcgca
     5521 ggccgcaggc cggaggcttt tccccagaga aaattaaaaa aattgatggg gcaaggccgc
     5581 aggccgcgca gttggagccg gtgggtatgt ggtcgaaggc tgggtagccg gtgggcaatc
     5641 cctgtggtca agctcgtggg caggcgcagc ctgtccatca gcttgtccag cagggttgtc
     5701 cacgggccga gcgaagcgag ccagccggtg gccgctcgcg gccatcgtcc acatatccac
     5761 gggctggcaa gggagcgcag cgaccgcgca gggcgaagcc cggagagcaa gcccgtaggg
     5821 ggggcgcgcc cagctgtcta gggcggcgga tttgtcctac tcaggagagc gttcaccgac
     5881 aaacacgaaa ggcccagtct ttcgactgag catttcgttt tatttgatgc ctttaattaa
     5941 acgttcgtaa tcaagccact tcctttttgc attgacgcag ggtgtcggaa ggcaactcgc
     6001 cgaacgcgct cctatagttt tcagcgaagc gtcccaaatg taagaagccg tagtctaggg
     6061 ctatctcagt tatactacgc acattggcac tgggatcgtt caagcaggcg cggatgcttt
     6121 cgagcttgcg gttgcggatg tagttcttcg gcgtggtgcc ggcgtgcttc tcgaacaaat
     6181 tgtagagcga gcgtggactc atcatcgcca gctccgctaa ccgctcaagg ctgatattcc
     6241 gtttgagatt ctcctcaatg aattgaacga ctcgctcgaa agacgggtta cctttgctga
     6301 aaatttcacg gctgacattg ctgcccagca tttcgagcag cttggaagcg atgatccccg
     6361 catagtgctc ttggacccga ggcatcgact ttgtatgttc cgcttcgtca caaactaacc
     6421 cgagtagatt gataaagcca tcgagttgct ggagattgtg tcgcgcggcg aaacggatac
     6481 cctccctcgg cttgtgccaa ttgttgtcac tgcacgcccg atcaaggacc actgagggca
     6541 atttaacgat aaatttctcg caatcttctg aataggtcag gtcggcttgg tcatccggat
     6601 tgagcagcaa tagttcgccc ggcgcaaaat agtgctcctg gccatggcca cgccacaggc
     6661 aatggccttt gagtattatt tgcagatgat aacaggtctc taatccaggc gagattaccc
     6721 tcacgctacc gccgtagctg attcgacaca gatcgaggca tccgaagatt ctgtggtgca
     6781 gcctgcctgc cgggcgcccg cccttgggca ggcgaataga gtgcgtaccg acatactggt
     6841 taacataatc ggagactgca tagggctcgg cgtggacgaa gatctgactt ttctcgttca
     6901 ataagcaaaa atccatagtt cacggttctc ttattttaat gtgggctgct tggtgtgatg
     6961 tagaaaggcg ccaagtcgat gaaaatgcat ctcgacgtga tgcgtatacg ggttaccccc
     7021 attgccacgt tgcgccatcc tttttgcaat cagtgaccac ttttccaagc aaaaataacg
     7081 ccaagcagaa cgaagacgtt ctttttaaga agcgagaaca ccagaagttc gtgctgtcgg
     7141 ggcatggggc gacgaattgg cggataaagg ggatctgctg gatattacgg cctttttaaa
     7201 gaccgtaaag aaaaataagc acaagtttta tccggccttt attcacattc ttgcccgcct
     7261 gatgaatgct catccgtaat tacgtatggc aatgaaagac ggtgagctgg tgatatggga
     7321 tagtgttcac ccttgttaca ccgttttcca tgagcaaact gaaacgtttt catcgctctg
     7381 gagtgaatac cacgacgatt tccggcagtt tctacacata tattcgcaag atgtggcgtg
     7441 ttacggtgaa aacctggcct atttccctaa agggtttatt gagaatatgt ttttcgtctc
     7501 agccaatccc tgggtgagtt tcaccagttt tgatttaaac gtggccaata tggacaactt
     7561 cttcgccccc gttttcacca tgcatgggaa tt
//
