LOCUS       sequence-393258-        6063 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the gene codying for the Ssr recombinase from Pseudomonas 
            putida DOT-T1E.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6063)
  AUTHORS   Pankratz D, Gomez NO, Nielsen A, Mustafayeva A, Gur M, 
            Arce-Rodriguez F, Nikel PI, Haussler S, Arce-Rodriguez A
  TITLE     An expanded CRISPR-Cas9-assisted recombineering toolkit for 
            engineering genetically intractable Pseudomonas aeruginosa isolates.
  JOURNAL   Nat Protoc. 2023 Oct 5. doi: 10.1038/s41596-023-00882-z.
  PUBMED    37798358
REFERENCE   2  (bases 1 to 6063)
  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: "Nat Protoc.
            2023 Oct 5. doi: 10.1038/s41596-023-00882-z."
COMMENT     Sequence Label: pSH824-ssr
FEATURES             Location/Qualifiers
     source          1..6063
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        11..88
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             89..1171
                     /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(108..127)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1403..1408,1409..1425,1426..1432)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1440..1456
                     /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)."
     terminator      2342..2436
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             2643..3443
                     /codon_start=1
                     /gene="aac(3)IV"
                     /product="aminoglycoside 3-N-acetyltransferase type IV     
                               "
                     /label=ApmR
                     /note="confers resistance to apramycin (Paget and Davies, 
                     1996)"
                     /translation="MSSAVECNVVQYEWRKAELIGQLLNLGVTPGGVLLVHSSFRSVRP
                     LEDGPLGLIEALRAALGPGGTLVMPSWSGLDDEPFDPATSPVTPDLGVVSDTFWRLPNV
                     KRSAHPFAFAAAGPQAEQIISDPLPLPPHSPASPVARVHELDGQVLLLGVGHDANTTLH
                     LAELMAKVPYGVPRHCTILQDGKLVRVDYLENDHCCERFALADRWLKEKSLQKEGPVGH
                     AFARLIRSRDIVATALGQLGRDPLIFLHPPEAGCEECDAARQSIG"
     oriT            3603..3711
                     /note="incP origin of transfer"
     CDS             complement(3772..4920)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     IVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     rep_origin      5313..5844
                     /label=oriV
                     /note="incP origin of replication"
     terminator      complement(5971..6057)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttaattaatt gacaccatcg aatggtgcaa aacctttcgc ggtatggcat gatagcgccc
       61 ggaagagagt caattcaggg tggtgaatgt gaaaccagta acgttatacg atgtcgcaga
      121 gtatgccggt gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc
      181 tgcgaaaacg cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg
      241 cgtggcacaa caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct
      301 ggccctgcac gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg
      361 tgccagcgtg gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt
      421 gcacaatctt ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca
      481 ggatgccatt gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgtctc
      541 tgaccagaca cccatcaaca gtattatttt ctcccatgaa gacggtacgc gactgggcgt
      601 ggagcatctg gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc cattaagttc
      661 tgtctcggcg cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca
      721 gccgatagcg gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca
      781 aatgctgaat gagggcatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct
      841 gggcgcaatg cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt
      901 gggatacgac gataccgaag acagctcatg ttatatcccg ccgttaacca ccatcaaaca
      961 ggattttcgc ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca
     1021 ggcggtgaag ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc
     1081 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     1141 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgta agttagcgcg
     1201 aattgatctg gtttgacagc ttatcatcga ctgcacggtg caccaatgct tctggcgtca
     1261 ggcagccatc ggaagctgtg gtatggctgt gcaggtcgta aatcactgca taattcgtgt
     1321 cgctcaaggc gcactcccgt tctggataat gttttttgcg ccgacatcat aacggttctg
     1381 gcaaatattc tgaaatgagc tgttgacaat taatcatccg gctcgtataa tgtgtggaat
     1441 tgtgagcgga taacaatttc acaccctagg ccgcggccgc gcgaattcag aaggagaata
     1501 taccatgagc caagtagcca gggtcgaaac ccattcccag ccacaggctg tagccgccga
     1561 gtcggcgacc atcctgcaga tcatccagca agtcgcaatg tcccccaatg cagacatcga
     1621 caagatggag cgattgatgg cgatgcaccg ccagcaccaa gcacaacagg cgcagcaagc
     1681 gttcgatgcc gcgctagccg ccatgcagga agagcttccg gttatccgtg agcgcggcgc
     1741 aatcaaggac aagtacaaga acgtccaatc tacctacgcc ctatgggagg acatcaacga
     1801 agagttgaag cccatcctgg cgaagcacgg ctttgctctc accttccgaa ttccacgcac
     1861 cgacaaaggc atcgaggttg agggcgtgct tagtcatcga gacggccaca gggaaaccac
     1921 atcaatcctc ctgccagccg atgctaccgg cagcaagaat gccgtccagg ccgttgccag
     1981 ttctgtcagc tacgggaagc gctacacggc cggcgctctt ctgaacttca ccactactgg
     2041 cgaagacgat gacggccaag gagccgtacc gatgcaagtg cctgatgagc cggtcatcac
     2101 accgcgccaa gccgcccaac tcgatgggct gctgaaaaaa tgcagccaag tcctggtcga
     2161 caacttcacc gccaagtacg gctgcgctgc caacgtctac aaatccgagt tcgacgttgt
     2221 gctcgctcgc ctcaccaagt cggccagccg gccgcaggag taaggatcct ctagagtcga
     2281 cctgcaggca tgcaagcttg cggccgcgtc gtgactggga aaaccctggc gactagtctt
     2341 ggactcctgt tgatagatcc agtaatgacc tcagaactcc atctggattt gttcagaacg
     2401 ctcggttgcc gccgggcgtt ttttattggt gagaatccag gggtccccaa taattacgat
     2461 ttaaatctca cggtaactga tgccgtattt gcagtaccag cgtacggccc acagaatgat
     2521 gtcacgctga aaatgccggc ctttgaatgg gttcatgtgc agctccatca gcaaaagggg
     2581 atgataagtt tatcaccacc gactatttgc aacagtgccg ttgatcgtgc tatgatcgag
     2641 tgatgtcatc agcggtggag tgcaatgtcg tgcaatacga atggcgaaaa gccgagctga
     2701 tcggtcagct tctcaacctt ggggttaccc ccggcggtgt gctgctggtc cacagctcct
     2761 tccgtagcgt ccggcccctc gaagatgggc cacttggact gatcgaggcc ctgcgtgctg
     2821 cgctgggtcc gggagggacg ctcgtgatgc cctcgtggtc aggtctggac gacgagccgt
     2881 tcgatcctgc cacgtcgccc gttacaccgg accttggagt tgtctctgac acattctggc
     2941 gcctgccaaa tgtaaagcgc agcgcccatc catttgcctt tgcggcagcg gggccacagg
     3001 cagagcagat catctctgat ccattgcccc tgccacctca ctcgcctgca agcccggtcg
     3061 cccgtgtcca tgaactcgat gggcaggtac ttctcctcgg cgtgggacac gatgccaaca
     3121 cgacgctgca tcttgccgag ttgatggcaa aggttcccta tggggtgccg agacactgca
     3181 ccattcttca ggatggcaag ttggttcgcg tcgattatct cgagaatgac cactgctgtg
     3241 agcgctttgc cttggcggac aggtggctca aggagaagag ccttcagaag gaaggtccag
     3301 tcggtcatgc ctttgctcgg ttgatccgct cccgcgacat tgtggcgaca gccctgggtc
     3361 aactgggccg agatccgttg atcttcctgc atccgccaga ggcgggatgc gaagaatgcg
     3421 atgccgctcg ccagtcgatt ggctgagacc gcggtccgcg cgttgtcctt ttccgctgca
     3481 taaccctgct tcggggtcat tatagcgatt ttttcggtat atccatcctt tttcgcacga
     3541 tatacaggat tttgccaaag ggttcgtgta gactttcctt ggtgtatcca acggcgtcag
     3601 ccgggcagga taggtgaagt aggcccaccc gcgagcgggt gttccttctt cactgtccct
     3661 tattcgcacc tggcggtgct caacgggaat cctgctctgc gaggctggcc gtaggccggc
     3721 cgcgatgcag gtggctgctg aacccccagc cggaactgac cccacaaggc cctagcgttt
     3781 gcaatgcacc aggtcatcat tgacccaggc gtgttccacc aggccgctgc ctcgcaactc
     3841 ttcgcaggct tcgccgacct gctcgcgcca cttcttcacg cgggtggaat ccgatccgca
     3901 catgaggcgg aaggtttcca gcttgagcgg gtacggctcc cggtgcgagc tgaaatagtc
     3961 gaacatccgt cgggccgtcg gcgacagctt gcggtacttc tcccatatga atttcgtgta
     4021 gtggtcgcca gcaaacagca cgacgatttc ctcgtcgatc aggacctggc aacgggacgt
     4081 tttcttgcca cggtccagga cgcggaagcg gtgcagcagc gacaccgatt ccaggtgccc
     4141 aacgcggtcg gacgtgaagc ccatcgccgt cgcctgtagg cgcgacaggc attcctcggc
     4201 cttcgtgtaa taccggccat tgatcgacca gcccaggtcc tggcaaagct cgtagaacgt
     4261 gaaggtgatc ggctcgccga taggggtgcg cttcgcgtac tccaacacct gctgccacac
     4321 cagttcgtca tcgtcggccc gcagctcgac gccggtgtag gtgatcttca cgtccttgtt
     4381 gacgtggaaa atgaccttgt tttgcagcgc ctcgcgcggg attttcttgt tgcgcgtggt
     4441 gaacagggca gagcgggccg tgtcgtttgg catcgctcgc atcgtgtccg gccacggcgc
     4501 aatatcgaac aaggaaagct gcatttcctt gatctgctgc ttcgtgtgtt tcagcaacgc
     4561 ggcctgcttg gcttcgctga cctgttttgc caggtcctcg ccggcggttt ttcgcttctt
     4621 ggtcgtcata gttcctcgcg tgtcgatggt catcgacttc gccaaacctg ccgcctcctg
     4681 ttcgagacga cgcgaacgct ccacggcggc cgatggcgcg ggcagggcag ggggagccag
     4741 ttgcacgctg tcgcgctcga tcttggccgt agcttgctgg actatcgagc cgacggactg
     4801 gaaggtttcg cggggcgcac gcatgacggt gcggcttgcg atggtttcgg catcctcggc
     4861 ggaaaacccc gcgtcgatca gttcttgcct gtatgccttc cggtcaaacg tccgattcat
     4921 tcaccctcct tgcgggattg ccccggaatt aattccccgg atcgatccgt cgatcttgat
     4981 cccctgcgcc atcagatcct tggcggcaag aaagccatcc agtttacttt gcagggcttc
     5041 ccaaccttac cagagggcgc cccagctggc aattccggtt cgcttgctgt ccataaaacc
     5101 gcccagtcta gctatcgcca tgtaagccca ctgcaagcta cctgctttct ctttgcgctt
     5161 gcgttttccc ttgtccagat agcccagtag ctgacattca tccggggtca gcaccgtttc
     5221 tgcggactgg ctttctacgt ggctgccatt tttggggtga ggccgttcgc ggccgagggg
     5281 cgcagcccct ggggggatgg gaggcccgcg ttagcgggcc gggagggttc gagaaggggg
     5341 ggcacccccc ttcggcgtgc gcggtcacgc gcacagggcg cagccctggt taaaaacaag
     5401 gtttataaat attggtttaa aagcaggtta aaagacaggt tagcggtggc cgaaaaacgg
     5461 gcggaaaccc ttgcaaatgc tggattttct gcctgtggac agcccctcaa atgtcaatag
     5521 gtgcgcccct catctgtcag cactctgccc ctcaagtgtc aaggatcgcg cccctcatct
     5581 gtcagtagtc gcgcccctca agtgtcaata ccgcagggca cttatcccca ggcttgtcca
     5641 catcatctgt gggaaactcg cgtaaaatca ggcgttttcg ccgatttgcg aggctggcca
     5701 gctccacgtc gccggccgaa atcgagcctg cccctcatct gtcaacgccg cgccgggtga
     5761 gtcggcccct caagtgtcaa cgtccgcccc tcatctgtca gtgagggcca agttttccgc
     5821 gaggtatcca caacgccggc ggccctacat ggctctgctg tagtgagtgg gttgcgctcc
     5881 ggcagcggtc ctgatccccc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     5941 tacggcgcgc ccagctgtct agggcggcgg atttgtccta ctcaggagag cgttcaccga
     6001 caaacaacag ataaaacgaa aggcccagtc tttcgactga gcctttcgtt ttatttgatg
     6061 cct
//
