LOCUS       sequence-408091-        3051 bp    DNA     circular SYN 14-APR-2025
DEFINITION  psrA repressor plasmid for LPattB2 integration.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3051)
  AUTHORS   Triassi AJ, Fields BD, Monahan CE, Means JM, Park Y, Doosthosseini 
            H, Padmakumar JP, Isabella VM, Voigt CA
  TITLE     Redesign of an Escherichia coli Nissle treatment for phenylketonuria
            using insulated genomic landing pads and genetic circuits to reduce 
            burden.
  JOURNAL   Cell Syst. 2023 Jun 21;14(6):512-524.e12. doi: 
            10.1016/j.cels.2023.05.004.
  PUBMED    37348465
REFERENCE   2  (bases 1 to 3051)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cels.2023.05"; journalName: "Cell Syst"; date: 
            "2023-06-21- 21"; volume: "14"; issue: "6"; pages: "512-524.e12"
COMMENT     Sequence Label: pAJT249-psrA
FEATURES             Location/Qualifiers
     source          1..3051
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      158..546
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     oriT            590..698
                     /note="incP origin of transfer"
     terminator      complement(849..943)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(954..1769)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(954..1769)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1677..1696
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(1770..1861)
                     /gene="bla"
                     /label=AmpR promoter
     protein_bind    1913..1960
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        join(2050..2055,2056..2071,2072..2078)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2084..2103
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
ORIGIN
        1 tcgcccgaga acttctgcaa ggcactgctc ttggctaggc ggtaatacgg ttatccacag
       61 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
      121 gtaaaaatga ttcgcacggg cccatggcta attcccatgt cagccgttaa gtgttcctgt
      181 gtcactcaaa attgctttga gaggctctaa gggcttctca gtgcgttaca tccctggctt
      241 gttgtccaca accgttaaac cttaaaagct ttaaaagcct tatatattct tttttttctt
      301 ataaaactta aaaccttaga ggctatttaa gttgctgatt tatattaatt ttattgttca
      361 aacatgagag cttagtacgt gaaacatgag agcttagtac gttagccatg agagcttagt
      421 acgttagcca tgagggttta gttcgttaaa catgagagct tagtacgtta aacatgagag
      481 cttagtacgt gaaacatgag agcttagtac gtactatcaa caggttgaac tgctgatctt
      541 cagatcctct acgccggacg catcgtggcc ggatcttctt gtagcttatc ggccagcctc
      601 gcagagcagg attcccgttg agcaccgcca ggtgcgaata agggacagtg aagaaggaac
      661 acccgctcgc gggtgggcct acttcaccta tcctgcccgg ctgacgccgt tggatacacc
      721 aaggaaagtc tacacgaacc ctttggcaaa atcctgtata tcgtgcgaaa aaggatggat
      781 ataccgaaaa aatcgctata atgaccccga agcagggtta tgcagcggaa agtatacctt
      841 aagcttggat tctcaccaat aaaaaacgcc cggcggcaac cgagcgttct gaacaaatcc
      901 agatggagtt ctgaggtcat tactggatct atcaacagga gtccaagcga gctttagaaa
      961 aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat
     1021 ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg
     1081 gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat
     1141 ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc
     1201 ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta
     1261 cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga
     1321 gcgagacgaa atacgcggtc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac
     1381 cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct
     1441 aatacctgga atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga
     1501 gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg
     1561 accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct
     1621 ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg
     1681 cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag
     1741 caagacgttt cccgttgaat atggctcata ctcttccttt ttcaatatta ttgaagcatt
     1801 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     1861 ataggcatgg tctaagaaac cattattatc atgacattaa cctataaaaa tagaagttcc
     1921 tattccgaag ttcctattct ctagaaagta taggaacttc ggatccgaat tcgagctccg
     1981 tcgacactag tgcggccgca ctcgagggca tcagagcaga ttgtactgac taaacgatcg
     2041 ttggctgtgt tgacaattaa tcatcggctc gtataatgtg tggaattgtg agcgctcaca
     2101 attagcgctg tctgtacttg tatcagtaca ctgacgagtc cctaaaggac gaaacaccgc
     2161 ctctacaaat aattttgttt aatttaattc gcggaagcgc agagataagg ggtatcatgg
     2221 cacagagcga aaccgttgaa cgtattctgg atgtagcaga acagctgttt gcagaacgtg
     2281 gttttgcaga aaccagcctg cgtctgatta ccagcaaagc cggtgttaat ctggcagcag
     2341 tgaattatca ttttggcagc aaaaaagcac tgattcaggc agtttttagc cgttttctgg
     2401 gtccgttttg tgcaagcctg gaacgtgaac tggaacgtcg tcaggcacgt ccggaacaga
     2461 aaccgagcct ggaagaactg ctggaaatgc tggttgaaca ggcactggca gttcagcctc
     2521 gtagcaataa tgatctgagc atttttatgt gtctgctggg tctggcattt agccagagcc
     2581 agggtcatct gcgtcgttat ctggaagata tgtatggtaa agtgttccgt cgttatatgc
     2641 tgctggttaa tgaagcagca ccgcgtgttc cgcctctgga actgttttgg cgtgttcatt
     2701 ttatgctggg tgcagcagca tttagcatga gcggtattaa agcactgcgt gcaattgcag
     2761 aaaccgattt tggtattaac accagcattg aacaggttat gcgtctgatg gttccgtttc
     2821 tggcagcagg tatgcgtgca gatagcggtg ttaccgatga agcaatggca gcagcacagc
     2881 tgcgtccgcg tagcaaaacc agcaccagcg caaccaccgc aaaagcataa cttattccat
     2941 aacaaagccg ggtaattccc ggctttgttg tatctgaaca ataaatggat gccctgcgta
     3001 agcggggcat ttttcttcct tgccgctcat gtatgtgtct acgcgagatt c
//
