LOCUS       sequence-243603-        4685 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  constitutive expression of I-SceI for genome engineering in 
            Pseudomonas.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4685)
  AUTHORS   Wirth NT, Kozaeva E, Nikel PI
  TITLE     Accelerated genome engineering of Pseudomonas putida by 
            I-SceI-mediated recombination and CRISPR-Cas9 counterselection.
  JOURNAL   Microb Biotechnol. 2019 Mar 12. doi: 10.1111/1751-7915.13396.
  PUBMED    30861315
REFERENCE   2  (bases 1 to 4685)
  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: "Microb 
            Biotechnol. 2019 Mar 12. doi: 10.1111/1751-7915.13396."
COMMENT     Sequence Label: pSEVA1213S
FEATURES             Location/Qualifiers
     source          1..4685
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    806..862
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     terminator      905..999
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     promoter        1072..1163
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1164..1232,1233..2024)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     terminator      2028..2067
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     oriT            2225..2333
                     /note="incP origin of transfer"
     CDS             complement(2394..3542)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIVSRTVMRAPRETFQSVGS
                     IVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     rep_origin      3935..4466
                     /label=oriV
                     /note="incP origin of replication"
     terminator      complement(4593..4679)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttaattaatg tgacaattaa tcatcggcat agtatatcgg catagtataa tacgacaagg
       61 tgaggaacta aacccctagg ggagtcatga acatatgaaa aacatcaaaa aaaaccaggt
      121 aatgaacctg ggtccgaact ctaaactgct gaaagaatac aaatcccagc tgatcgaact
      181 gaacatcgaa cagttcgaag caggtatcgg tctgatcctg ggtgatgctt acatccgttc
      241 tcgtgatgaa ggtaaaacct actgtatgca gttcgagtgg aaaaacaaag catacatgga
      301 ccacgtatgt ctgctgtacg atcagtgggt actgtccccg ccgcacaaaa aagaacgtgt
      361 taaccacctg ggtaacctgg taatcacctg gggcgcccag actttcaaac accaggcttt
      421 caacaaactg gctaacctgt tcatcgttaa caacaaaaaa accatcccga acaacctggt
      481 tgaaaactac ctgaccccga tgtctctggc atactggttc atggatgatg gtggtaaatg
      541 ggattacaac aaaaactcta ccaacaaatc gatcgtactg aacacccagt ctttcacttt
      601 cgaagaagta gaatacctgg ttaagggtct gcgtaacaaa ttccaactga actgttacgt
      661 aaaaatcaac aaaaacaaac cgatcatcta catcgattct atgtcttacc tgatcttcta
      721 caacctgatc aaaccgtacc tgatcccgca gatgatgtac aaactgccga acactatctc
      781 ctccgaaact ttcctgaaat aataagaatt cgagctcggt acccggggat cctctagagt
      841 cgacctgcag gcatgcaagc ttgcggccgc gtcgtgactg ggaaaaccct ggcgactagt
      901 cttggactcc tgttgataga tccagtaatg acctcagaac tccatctgga tttgttcaga
      961 acgctcggtt gccgccgggc gttttttatt ggtgagaatc caggggtccc caataattac
     1021 gatttaaatt agtagcccgc ctaatgagcg ggcttttttt taattcccct atttgtttat
     1081 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     1141 aataatattg aaaaaggaag agtatgagca ttcagcattt tcgtgtggcg ctgattccgt
     1201 tttttgcggc gttttgcctg ccggtgtttg cgcatccgga aaccctggtg aaagtgaaag
     1261 atgcggaaga tcaactgggt gcgcgcgtgg gctatattga actggatctg aacagcggca
     1321 aaattctgga atcttttcgt ccggaagaac gttttccgat gatgagcacc tttaaagtgc
     1381 tgctgtgcgg tgcggttctg agccgtgtgg atgcgggcca ggaacaactg ggccgtcgta
     1441 ttcattatag ccagaacgat ctggtggaat atagcccggt gaccgaaaaa catctgaccg
     1501 atggcatgac cgtgcgtgaa ctgtgcagcg cggcgattac catgagcgat aacaccgcgg
     1561 cgaacctgct gctgacgacc attggcggtc cgaaagaact gaccgcgttt ctgcataaca
     1621 tgggcgatca tgtgacccgt ctggatcgtt gggaaccgga actgaacgaa gcgattccga
     1681 acgatgaacg tgataccacc atgccggcag caatggcgac caccctgcgt aaactgctga
     1741 cgggtgagct gctgaccctg gcaagccgcc agcaactgat tgattggatg gaagcggata
     1801 aagtggcggg tccgctgctg cgtagcgcgc tgccggctgg ctggtttatt gcggataaaa
     1861 gcggtgcggg cgaacgtggc agccgtggca ttattgcggc gctgggcccg gatggtaaac
     1921 cgagccgtat tgtggtgatt tataccaccg gcagccaggc gacgatggat gaacgtaacc
     1981 gtcagattgc ggaaattggc gcgagcctga ttaaacattg gtaaaccgat acaattaaag
     2041 gctccttttg gagccttttt ttttggacga ccccggtccg cgcgttgtcc ttttccgctg
     2101 cataaccctg cttcggggtc attatagcga ttttttcggt atatccatcc tttttcgcac
     2161 gatatacagg attttgccaa agggttcgtg tagactttcc ttggtgtatc caacggcgtc
     2221 agccgggcag gataggtgaa gtaggcccac ccgcgagcgg gtgttccttc ttcactgtcc
     2281 cttattcgca cctggcggtg ctcaacggga atcctgctct gcgaggctgg ccgtaggccg
     2341 gccgcgatgc aggtggctgc tgaaccccca gccggaactg accccacaag gccctagcgt
     2401 ttgcaatgca ccaggtcatc attgacccag gcgtgttcca ccaggccgct gcctcgcaac
     2461 tcttcgcagg cttcgccgac ctgctcgcgc cacttcttca cgcgggtgga atccgatccg
     2521 cacatgaggc ggaaggtttc cagcttgagc gggtacggct cccggtgcga gctgaaatag
     2581 tcgaacatcc gtcgggccgt cggcgacagc ttgcggtact tctcccatat gaatttcgtg
     2641 tagtggtcgc cagcaaacag cacgacgatt tcctcgtcga tcaggacctg gcaacgggac
     2701 gttttcttgc cacggtccag gacgcggaag cggtgcagca gcgacaccga ttccaggtgc
     2761 ccaacgcggt cggacgtgaa gcccatcgcc gtcgcctgta ggcgcgacag gcattcctcg
     2821 gccttcgtgt aataccggcc attgatcgac cagcccaggt cctggcaaag ctcgtagaac
     2881 gtgaaggtga tcggctcgcc gataggggtg cgcttcgcgt actccaacac ctgctgccac
     2941 accagttcgt catcgtcggc ccgcagctcg acgccggtgt aggtgatctt cacgtccttg
     3001 ttgacgtgga aaatgacctt gttttgcagc gcctcgcgcg ggattttctt gttgcgcgtg
     3061 gtgaacaggg cagagcgggc cgtgtcgttt ggcatcgctc gcatcgtgtc cggccacggc
     3121 gcaatatcga acaaggaaag ctgcatttcc ttgatctgct gcttcgtgtg tttcagcaac
     3181 gcggcctgct tggcttcgct gacctgtttt gccaggtcct cgccggcggt ttttcgcttc
     3241 ttggtcgtca tagttcctcg cgtgtcgatg gtcatcgact tcgccaaacc tgccgcctcc
     3301 tgttcgagac gacgcgaacg ctccacggcg gccgatggcg cgggcagggc agggggagcc
     3361 agttgcacgc tgtcgcgctc gatcttggcc gtagcttgct ggactatcga gccgacggac
     3421 tggaaggttt cgcggggcgc acgcatgacg gtgcggctta cgatggtttc ggcatcctcg
     3481 gcggaaaacc ccgcgtcgat cagttcttgc ctgtatgcct tccggtcaaa cgtccgattc
     3541 attcaccctc cttgcgggat tgccccggaa ttaattcccc ggatcgatcc gtcgatcttg
     3601 atcccctgcg ccatcagatc cttggcggca agaaagccat ccagtttact ttgcagggct
     3661 tcccaacctt accagagggc gccccagctg gcaattccgg ttcgcttgct gtccataaaa
     3721 ccgcccagtc tagctatcgc catgtaagcc cactgcaagc tacctgcttt ctctttgcgc
     3781 ttgcgttttc ccttgtccag atagcccagt agctgacatt catccggggt cagcaccgtt
     3841 tctgcggact ggctttctac gtggctgcca tttttggggt gaggccgttc gcggccgagg
     3901 ggcgcagccc ctggggggat gggaggcccg cgttagcggg ccgggagggt tcgagaaggg
     3961 ggggcacccc ccttcggcgt gcgcggtcac gcgcacaggg cgcagccctg gttaaaaaca
     4021 aggtttataa atattggttt aaaagcaggt taaaagacag gttagcggtg gccgaaaaac
     4081 gggcggaaac ccttgcaaat gctggatttt ctgcctgtgg acagcccctc aaatgtcaat
     4141 aggtgcgccc ctcatctgtc agcactctgc ccctcaagtg tcaaggatcg cgcccctcat
     4201 ctgtcagtag tcgcgcccct caagtgtcaa taccgcaggg cacttatccc caggcttgtc
     4261 cacatcatct gtgggaaact cgcgtaaaat caggcgtttt cgccgatttg cgaggctggc
     4321 cagctccacg tcgccggccg aaatcgagcc tgcccctcat ctgtcaacgc cgcgccgggt
     4381 gagtcggccc ctcaagtgtc aacgtccgcc cctcatctgt cagtgagggc caagttttcc
     4441 gcgaggtatc cacaacgccg gcggccctac atggctctgc tgtagtgagt gggttgcgct
     4501 ccggcagcgg tcctgatccc ccgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4561 tctacggcgc gcccagctgt ctagggcggc ggatttgtcc tactcaggag agcgttcacc
     4621 gacaaacaac agataaaacg aaaggcccag tctttcgact gagcctttcg ttttatttga
     4681 tgcct
//
