LOCUS       sequence-408078-        2986 bp    DNA     circular SYN 14-APR-2025
DEFINITION  RPU integration plasmid (LPattB7).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2986)
  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 2986)
  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: pAJT430
FEATURES             Location/Qualifiers
     source          1..2986
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      265..653
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     oriT            697..805
                     /note="incP origin of transfer"
     terminator      complement(956..1049)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(1060..1719)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     NTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYDENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1634..1653
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(1720..1822)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     protein_bind    1939..1986
                     /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        2088..2122
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     misc_RNA        2129..2178
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     RBS             2210..2221
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             join(2228..2230,2231..2233,2234..2947)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2273..2294)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2534..2553)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2881..2902
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 cctaacctta acttttacgc aggttcagct tggaaacaca gaaaaaagcc cgcacctgac
       61 agtgcgggct ttttttttcg accaaaggct cggtaccaaa ttccagaaaa gacacccgaa
      121 agggtgtttt ttcgttttgg tccaggcggt aatacggtta tccacagaat ccggggataa
      181 cgcaggaaag aacatgttag caaaaggcca gcaaaaggcc aggaaccgta aaaatgattc
      241 gcacgggccc atggctaatt cccatgtcag ccgttaagtg ttcctgtgtc actcaaaatt
      301 gctttgagag gctctaaggg cttctcagtg cgttacatcc ctggcttgtt gtccacaacc
      361 gttaaacctt aaaagcttta aaagccttat atattctttt ttttcttata aaacttaaaa
      421 ccttagaggc tatttaagtt gctgatttat attaatttta ttgttcaaac atgagagctt
      481 agtacgtgaa acatgagagc ttagtacgtt agccatgaga gcttagtacg ttagccatga
      541 gggtttagtt cgttaaacat gagagcttag tacgttaaac atgagagctt agtacgtgaa
      601 acatgagagc ttagtacgta ctatcaacag gttgaactgc tgatcttcag atcctctacg
      661 ccggacgcat cgtggccgga tcttcttgta gcttatcggc cagcctcgca gagcaggatt
      721 cccgttgagc accgccaggt gcgaataagg gacagtgaag aaggaacacc cgctcgcggg
      781 tgggcctact tcacctatcc tgcccggctg acgccgttgg atacaccaag gaaagtctac
      841 acgaaccctt tggcaaaatc ctgtatatcg tgcgaaaaag gatggatata ccgaaaaaat
      901 cgctataatg accccgaagc agggttatgc agcggaaagt ataccttaag cttggattct
      961 caccataaaa aacgcccggc ggcaaccgag cgttctgaac aaatccagat ggagttctga
     1021 ggtcattact ggatctatca acaggagtcc aagcgagctt tacgccccgc cctgccactc
     1081 atcgcagtac tgttgtaatt cattaagcat tctgccgaca tggaagccat cacaaacggc
     1141 atgatgaacc tgaatcgcca gcggcatcag caccttgtcg ccttgcgtat aatatttgcc
     1201 catggtgaaa acgggggcga agaagttgtc catattggcc acgtttaaat caaaactggt
     1261 gaaactcacc cagggattgg ctgagacgaa aaacatattc tcaataaacc ctttagggaa
     1321 ataggccagg ttttcatcgt aacacgccac atcttgcgaa tatatgtgta gaaactgccg
     1381 gaaatcgtcg tggtattcac tccagagcga tgaaaacgtt tcagtttgct catggaaaac
     1441 ggtgtaacaa gggtgaacac tatcccatat caccagctca ccgtctttca ttgccatacg
     1501 aaattccgga tgagcattca tcaggcgggc aagaatgtga ataaaggccg gataaaactt
     1561 gtgcttattt ttctttacgg tatttaaaaa ggccgtaata tccagctgaa cggtctggtt
     1621 ataggtacat tgagcaactg actgaaatgc ctcaaaatgt tctttacgat gccattggga
     1681 tatatcaacg gtggtatatc cagtgatttt tttctccatt ttagcttcct tagctcctga
     1741 aaatctcgat aactcaaaaa atacgcccgg tagtgatctt atttcattat ggtgaaagtt
     1801 ggaacctctt acgtgccgat caacgtctca ttttcgccaa tttaaatcgt aattattggg
     1861 gaccctgtat ttagaaaaat aaacaaatag gcatggtcta agaaaccatt attatcatga
     1921 cattaaccta taaaaataga agttcctatt ccgaagttcc tattctctag aaagtatagg
     1981 aacttcggat ccgaattcga gctccgtcga cactagtgcg gccgcactcg agggcatcag
     2041 agcagattgg gagcgacgta cggtggaagc ttgataagtc cctaactttt acagctagct
     2101 cagtcctagg tattatgcta gcctgaagct gtcaccggat gtgctttccg gtctgatgag
     2161 tccgtgagga cgaaacagcc tctacaaata attttgttta atactagaga aagaggagaa
     2221 atactagatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     2281 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     2341 cacctacggc aagctgaccc tgaagttcat ctgcaccaca ggcaagctgc ccgtgccctg
     2401 gcccaccctc gtgaccacct tcggctacgg cctgcaatgc ttcgcccgct accccgacca
     2461 catgaagctg cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     2521 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     2581 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     2641 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca
     2701 gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     2761 gctcgccgac cactaccagc agaacacccc aatcggcgac ggccccgtgc tgctgcccga
     2821 caaccactac cttagctacc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     2881 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     2941 caagtaagct tgtgttataa acctgtgtga gagttaagtt tacatg
//
