LOCUS       sequence-393257-        5888 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 5888)
  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 5888)
  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: pSH624-ssr
FEATURES             Location/Qualifiers
     source          1..5888
                     /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"
     promoter        join(2467..2472,2473..2489,2490..2495)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             2684..3217
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     oriT            3428..3536
                     /note="incP origin of transfer"
     CDS             complement(3597..4745)
                     /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      5138..5669
                     /label=oriV
                     /note="incP origin of replication"
     terminator      complement(5796..5882)
                     /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 ttaaatttga cataagcctg ttcggttcgt aaactgtaat gcaagtagcg tatgcgctca
     2521 cgcaactggt ccagaacctt gaccgaacgc agcggtggta acggcgcagt ggcggttttc
     2581 atggcttgtt atgactgttt ttttgtacag cctatgcctc gggcatccaa gcagcaagcg
     2641 cgttacgccg tgggtcgatg tttgatgtta tggagcagca acgatgttac gcagcagcaa
     2701 cgatgttacg cagcagggca gtcgccctaa aacaaagtta ggtggctcaa gtatgggcat
     2761 cattcgcaca tgtaggctcg gccctgacca agtcaaatcc atgcgggctg ctcttgatct
     2821 tttcggtcgt gagttcggag acgtagccac ctactcccaa catcagccgg actccgatta
     2881 cctcgggaac ttgctccgta gtaagacatt catcgcgctt gctgccttcg accaagaagc
     2941 ggttgttggc gctctcgcgg cttacgttct gcccaagttt gagcagccgc gtagtgagat
     3001 ctatatctat gatctcgcag tctccggaga gcaccggagg cagggcattg ccaccgcgct
     3061 catcaatctc ctcaagcatg aggccaacgc gcttggtgct tatgtgatct acgtgcaagc
     3121 agattacggt gacgatcccg cagtggctct ctatacaaag ttgggcatac gggaagaagt
     3181 gatgcacttt gatatcgacc caagtaccgc cacctaacaa ttcgttcaag ccgagatcgg
     3241 cttcccggcc gcggagttgt tcggtaaatt ggacaacggt ccgcgcgttg tccttttccg
     3301 ctgcataacc ctgcttcggg gtcattatag cgattttttc ggtatatcca tcctttttcg
     3361 cacgatatac aggattttgc caaagggttc gtgtagactt tccttggtgt atccaacggc
     3421 gtcagccggg caggataggt gaagtaggcc cacccgcgag cgggtgttcc ttcttcactg
     3481 tcccttattc gcacctggcg gtgctcaacg ggaatcctgc tctgcgaggc tggccgtagg
     3541 ccggccgcga tgcaggtggc tgctgaaccc ccagccggaa ctgaccccac aaggccctag
     3601 cgtttgcaat gcaccaggtc atcattgacc caggcgtgtt ccaccaggcc gctgcctcgc
     3661 aactcttcgc aggcttcgcc gacctgctcg cgccacttct tcacgcgggt ggaatccgat
     3721 ccgcacatga ggcggaaggt ttccagcttg agcgggtacg gctcccggtg cgagctgaaa
     3781 tagtcgaaca tccgtcgggc cgtcggcgac agcttgcggt acttctccca tatgaatttc
     3841 gtgtagtggt cgccagcaaa cagcacgacg atttcctcgt cgatcaggac ctggcaacgg
     3901 gacgttttct tgccacggtc caggacgcgg aagcggtgca gcagcgacac cgattccagg
     3961 tgcccaacgc ggtcggacgt gaagcccatc gccgtcgcct gtaggcgcga caggcattcc
     4021 tcggccttcg tgtaataccg gccattgatc gaccagccca ggtcctggca aagctcgtag
     4081 aacgtgaagg tgatcggctc gccgataggg gtgcgcttcg cgtactccaa cacctgctgc
     4141 cacaccagtt cgtcatcgtc ggcccgcagc tcgacgccgg tgtaggtgat cttcacgtcc
     4201 ttgttgacgt ggaaaatgac cttgttttgc agcgcctcgc gcgggatttt cttgttgcgc
     4261 gtggtgaaca gggcagagcg ggccgtgtcg tttggcatcg ctcgcatcgt gtccggccac
     4321 ggcgcaatat cgaacaagga aagctgcatt tccttgatct gctgcttcgt gtgtttcagc
     4381 aacgcggcct gcttggcttc gctgacctgt tttgccaggt cctcgccggc ggtttttcgc
     4441 ttcttggtcg tcatagttcc tcgcgtgtcg atggtcatcg acttcgccaa acctgccgcc
     4501 tcctgttcga gacgacgcga acgctccacg gcggccgatg gcgcgggcag ggcaggggga
     4561 gccagttgca cgctgtcgcg ctcgatcttg gccgtagctt gctggactat cgagccgacg
     4621 gactggaagg tttcgcgggg cgcacgcatg acggtgcggc ttgcgatggt ttcggcatcc
     4681 tcggcggaaa accccgcgtc gatcagttct tgcctgtatg ccttccggtc aaacgtccga
     4741 ttcattcacc ctccttgcgg gattgccccg gaattaattc cccggatcga tccgtcgatc
     4801 ttgatcccct gcgccatcag atccttggcg gcaagaaagc catccagttt actttgcagg
     4861 gcttcccaac cttaccagag ggcgccccag ctggcaattc cggttcgctt gctgtccata
     4921 aaaccgccca gtctagctat cgccatgtaa gcccactgca agctacctgc tttctctttg
     4981 cgcttgcgtt ttcccttgtc cagatagccc agtagctgac attcatccgg ggtcagcacc
     5041 gtttctgcgg actggctttc tacgtggctg ccatttttgg ggtgaggccg ttcgcggccg
     5101 aggggcgcag cccctggggg gatgggaggc ccgcgttagc gggccgggag ggttcgagaa
     5161 gggggggcac cccccttcgg cgtgcgcggt cacgcgcaca gggcgcagcc ctggttaaaa
     5221 acaaggttta taaatattgg tttaaaagca ggttaaaaga caggttagcg gtggccgaaa
     5281 aacgggcgga aacccttgca aatgctggat tttctgcctg tggacagccc ctcaaatgtc
     5341 aataggtgcg cccctcatct gtcagcactc tgcccctcaa gtgtcaagga tcgcgcccct
     5401 catctgtcag tagtcgcgcc cctcaagtgt caataccgca gggcacttat ccccaggctt
     5461 gtccacatca tctgtgggaa actcgcgtaa aatcaggcgt tttcgccgat ttgcgaggct
     5521 ggccagctcc acgtcgccgg ccgaaatcga gcctgcccct catctgtcaa cgccgcgccg
     5581 ggtgagtcgg cccctcaagt gtcaacgtcc gcccctcatc tgtcagtgag ggccaagttt
     5641 tccgcgaggt atccacaacg ccggcggccc tacatggctc tgctgtagtg agtgggttgc
     5701 gctccggcag cggtcctgat cccccgcaga aaaaaaggat ctcaagaaga tcctttgatc
     5761 ttttctacgg cgcgcccagc tgtctagggc ggcggatttg tcctactcag gagagcgttc
     5821 accgacaaac aacagataaa acgaaaggcc cagtctttcg actgagcctt tcgttttatt
     5881 tgatgcct
//
