LOCUS       sequence-393261-        6080 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 6080)
  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 6080)
  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: pSH424-ssr
FEATURES             Location/Qualifiers
     source          1..6080
                     /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             2672..3463
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     oriT            3620..3728
                     /note="incP origin of transfer"
     CDS             complement(3789..4937)
                     /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      5330..5861
                     /label=oriV
                     /note="incP origin of replication"
     terminator      complement(5988..6074)
                     /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 ttacgtattt aaatgaacct tgaccgaacg cagcggtggt aacggcgcag tggcggtttt
     2521 catggcttgt tatgactgtt tttttggggt acagtctatg cctcgggcat ccaagcagca
     2581 agcgcgttac gccgtgggtc gatgtttgat gttatggagc agcaacgatg ttacgcagca
     2641 gggcagtcgc cctaaaacaa agttaaacat catgagggaa gcggtgatcg ccgaagtatc
     2701 gactcaacta tcagaggtag ttggcgtcat cgagcgccat ctcgaaccga cgttgctggc
     2761 cgtacatttg tacggctccg cagtggatgg cggcctgaag ccacacagtg atattgattt
     2821 gctggttacg gtgaccgtaa ggcttgatga aacaacgcgg cgagctttga tcaacgacct
     2881 tttggaaact tcggcttccc ctggagagag cgagattctc cgcgctgtag aagtcaccat
     2941 tgttgtgcac gacgacatca ttccgtggcg ttatccagct aagcgcgaac tgcaatttgg
     3001 agaatggcag cgcaatgaca ttcttgcagg tatcttcgag ccagccacga tcgacattga
     3061 tctggctatc ttgctgacaa aagcaagaga acatagcgtt gccttggtag gtccagcggc
     3121 ggaggaactc tttgatccgg ttcctgaaca ggatctattt gaggcgctaa atgaaacctt
     3181 aacgctatgg aactcgccgc ccgactgggc tggcgatgag cgaaatgtag tgcttacgtt
     3241 gtcccgcatt tggtacagcg cagtaaccgg caaaatcgcg ccgaaggatg tcgctgccga
     3301 ctgggcaatg gagcgcctgc cggcccagta tcagcccgtc atacttgaag ctagacaggc
     3361 ttatcttgga caagaagaag atcgcttggc ctcgcgcgca gatcagttgg aagaatttgt
     3421 ccactacgtg aaaggcgaga tcaccaaggt agtcggcaaa taagacaacg gtccgcgcgt
     3481 tgtccttttc cgctgcataa ccctgcttcg gggtcattat agcgattttt tcggtatatc
     3541 catccttttt cgcacgatat acaggatttt gccaaagggt tcgtgtagac tttccttggt
     3601 gtatccaacg gcgtcagccg ggcaggatag gtgaagtagg cccacccgcg agcgggtgtt
     3661 ccttcttcac tgtcccttat tcgcacctgg cggtgctcaa cgggaatcct gctctgcgag
     3721 gctggccgta ggccggccgc gatgcaggtg gctgctgaac ccccagccgg aactgacccc
     3781 acaaggccct agcgtttgca atgcaccagg tcatcattga cccaggcgtg ttccaccagg
     3841 ccgctgcctc gcaactcttc gcaggcttcg ccgacctgct cgcgccactt cttcacgcgg
     3901 gtggaatccg atccgcacat gaggcggaag gtttccagct tgagcgggta cggctcccgg
     3961 tgcgagctga aatagtcgaa catccgtcgg gccgtcggcg acagcttgcg gtacttctcc
     4021 catatgaatt tcgtgtagtg gtcgccagca aacagcacga cgatttcctc gtcgatcagg
     4081 acctggcaac gggacgtttt cttgccacgg tccaggacgc ggaagcggtg cagcagcgac
     4141 accgattcca ggtgcccaac gcggtcggac gtgaagccca tcgccgtcgc ctgtaggcgc
     4201 gacaggcatt cctcggcctt cgtgtaatac cggccattga tcgaccagcc caggtcctgg
     4261 caaagctcgt agaacgtgaa ggtgatcggc tcgccgatag gggtgcgctt cgcgtactcc
     4321 aacacctgct gccacaccag ttcgtcatcg tcggcccgca gctcgacgcc ggtgtaggtg
     4381 atcttcacgt ccttgttgac gtggaaaatg accttgtttt gcagcgcctc gcgcgggatt
     4441 ttcttgttgc gcgtggtgaa cagggcagag cgggccgtgt cgtttggcat cgctcgcatc
     4501 gtgtccggcc acggcgcaat atcgaacaag gaaagctgca tttccttgat ctgctgcttc
     4561 gtgtgtttca gcaacgcggc ctgcttggct tcgctgacct gttttgccag gtcctcgccg
     4621 gcggtttttc gcttcttggt cgtcatagtt cctcgcgtgt cgatggtcat cgacttcgcc
     4681 aaacctgccg cctcctgttc gagacgacgc gaacgctcca cggcggccga tggcgcgggc
     4741 agggcagggg gagccagttg cacgctgtcg cgctcgatct tggccgtagc ttgctggact
     4801 atcgagccga cggactggaa ggtttcgcgg ggcgcacgca tgacggtgcg gcttgcgatg
     4861 gtttcggcat cctcggcgga aaaccccgcg tcgatcagtt cttgcctgta tgccttccgg
     4921 tcaaacgtcc gattcattca ccctccttgc gggattgccc cggaattaat tccccggatc
     4981 gatccgtcga tcttgatccc ctgcgccatc agatccttgg cggcaagaaa gccatccagt
     5041 ttactttgca gggcttccca accttaccag agggcgcccc agctggcaat tccggttcgc
     5101 ttgctgtcca taaaaccgcc cagtctagct atcgccatgt aagcccactg caagctacct
     5161 gctttctctt tgcgcttgcg ttttcccttg tccagatagc ccagtagctg acattcatcc
     5221 ggggtcagca ccgtttctgc ggactggctt tctacgtggc tgccattttt ggggtgaggc
     5281 cgttcgcggc cgaggggcgc agcccctggg gggatgggag gcccgcgtta gcgggccggg
     5341 agggttcgag aagggggggc accccccttc ggcgtgcgcg gtcacgcgca cagggcgcag
     5401 ccctggttaa aaacaaggtt tataaatatt ggtttaaaag caggttaaaa gacaggttag
     5461 cggtggccga aaaacgggcg gaaacccttg caaatgctgg attttctgcc tgtggacagc
     5521 ccctcaaatg tcaataggtg cgcccctcat ctgtcagcac tctgcccctc aagtgtcaag
     5581 gatcgcgccc ctcatctgtc agtagtcgcg cccctcaagt gtcaataccg cagggcactt
     5641 atccccaggc ttgtccacat catctgtggg aaactcgcgt aaaatcaggc gttttcgccg
     5701 atttgcgagg ctggccagct ccacgtcgcc ggccgaaatc gagcctgccc ctcatctgtc
     5761 aacgccgcgc cgggtgagtc ggcccctcaa gtgtcaacgt ccgcccctca tctgtcagtg
     5821 agggccaagt tttccgcgag gtatccacaa cgccggcggc cctacatggc tctgctgtag
     5881 tgagtgggtt gcgctccggc agcggtcctg atcccccgca gaaaaaaagg atctcaagaa
     5941 gatcctttga tcttttctac ggcgcgccca gctgtctagg gcggcggatt tgtcctactc
     6001 aggagagcgt tcaccgacaa acaacagata aaacgaaagg cccagtcttt cgactgagcc
     6061 tttcgtttta tttgatgcct
//
