LOCUS       sequence-408087-        3241 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PCin-YFP plasmid for LPattB2 integration.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3241)
  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 3241)
  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"; 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: pAJT243-Pcin
FEATURES             Location/Qualifiers
     source          1..3241
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(28..741,742..744,745..747))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(73..94)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     422..441
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     681..702
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     RBS             754..765
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     misc_RNA        complement(797..846)
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     protein_bind    complement(1170..1217)
                     /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        1269..1360
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1361..2176
                     /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             1361..2176
                     /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     complement(1434..1453)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2187..2281
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     oriT            2432..2540
                     /note="incP origin of transfer"
     rep_origin      2584..2972
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
ORIGIN
        1 gaatctcgcg tagacacata catgagctta cttgtacagc tcgtccatgc cgagagtgat
       61 cccggcggcg gtcacgaact ccagcaggac catgtgatcg cgcttctcgt tggggtcttt
      121 gctcagggcg gactggtagc taaggtagtg gttgtcgggc agcagcacgg ggccgtcgcc
      181 gattggggtg ttctgctggt agtggtcggc gagctgcacg ctgccgtcct cgatgttgtg
      241 gcggatcttg aagttcacct tgatgccgtt cttctgcttg tcggccatga tatagacgtt
      301 gtggctgttg tagttgtact ccagcttgtg ccccaggatg ttgccgtcct ccttgaagtc
      361 gatgcccttc agctcgatgc ggttcaccag ggtgtcgccc tcgaacttca cctcggcgcg
      421 ggtcttgtag ttgccgtcgt ccttgaagaa gatggtgcgc tcctggacgt agccttcggg
      481 catggcggac ttgaagaagt cgtgcagctt catgtggtcg gggtagcggg cgaagcattg
      541 caggccgtag ccgaaggtgg tcacgagggt gggccagggc acgggcagct tgcctgtggt
      601 gcagatgaac ttcagggtca gcttgccgta ggtggcatcg ccctcgccct cgccggacac
      661 gctgaacttg tggccgttta cgtcgccgtc cagctcgacc aggatgggca ccaccccggt
      721 gaacagctcc tcgcccttgc tcaccatcta gtatttctcc tctttctcta gtattaaaca
      781 aaattatttg tagaggctgt ttcgtcctca cggactcatc agaccggaaa gcacatccgg
      841 tgacagctag cgttttcaag ttcgtggaaa gcagaatggt tcaggcgagc cggatcggaa
      901 attccctcag ataggccccc ccagatgagg gggatcagag catgtcgcgc aaaagtgtgc
      961 agcggttttg cgataacgac atgcgccaaa acaaagactg aaagcgcgag gagcgaatct
     1021 gaaagatcgc gacccgcttt agagcgccgt gcgtccgttt ggacgcacaa agggactttt
     1081 gtgctaccac tcctggtctt ccagtacaat ctgctctgat gccctcgagt gcggccgcac
     1141 tagtgtcgac ggagctcgaa ttcggatccg aagttcctat actttctaga gaataggaac
     1201 ttcggaatag gaacttctat ttttataggt taatgtcatg ataataatgg tttcttagac
     1261 catgcctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     1321 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagccata ttcaacggga
     1381 aacgtcttgc tctaggccgc gattaaattc caacatggat gctgatttat atgggtataa
     1441 atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc tatcgattgt atgggaagcc
     1501 cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc gttgccaatg atgttacaga
     1561 tgagatggtc agactaaact ggctgacgga atttatgcct cttccgacca tcaagcattt
     1621 tatccgtact cctgatgatg catggttact caccactgcg atccccggga aaacagcatt
     1681 ccaggtatta gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc tggcagtgtt
     1741 cctgcgccgg ttgcattcga ttcctgtttg taattgtcct tttaacagcg accgcgtatt
     1801 tcgtctcgct caggcgcaat cacgaatgaa taacggtttg gttgatgcga gtgattttga
     1861 tgacgagcgt aatggctggc ctgttgaaca agtctggaaa gaaatgcata aacttttgcc
     1921 attctcaccg gattcagtcg tcactcatgg tgatttctca cttgataacc ttatttttga
     1981 cgaggggaaa ttaataggtt gtattgatgt tggacgagtc ggaatcgcag accgatacca
     2041 ggatcttgcc atcctatgga actgcctcgg tgagttttct ccttcattac agaaacggct
     2101 ttttcaaaaa tatggtattg ataatcctga tatgaataaa ttgcagtttc atttgatgct
     2161 cgatgagttt ttctaaagct cgcttggact cctgttgata gatccagtaa tgacctcaga
     2221 actccatctg gatttgttca gaacgctcgg ttgccgccgg gcgtttttta ttggtgagaa
     2281 tccaagctta aggtatactt tccgctgcat aaccctgctt cggggtcatt atagcgattt
     2341 tttcggtata tccatccttt ttcgcacgat atacaggatt ttgccaaagg gttcgtgtag
     2401 actttccttg gtgtatccaa cggcgtcagc cgggcaggat aggtgaagta ggcccacccg
     2461 cgagcgggtg ttccttcttc actgtccctt attcgcacct ggcggtgctc aacgggaatc
     2521 ctgctctgcg aggctggccg ataagctaca agaagatccg gccacgatgc gtccggcgta
     2581 gaggatctga agatcagcag ttcaacctgt tgatagtacg tactaagctc tcatgtttca
     2641 cgtactaagc tctcatgttt aacgtactaa gctctcatgt ttaacgaact aaaccctcat
     2701 ggctaacgta ctaagctctc atggctaacg tactaagctc tcatgtttca cgtactaagc
     2761 tctcatgttt gaacaataaa attaatataa atcagcaact taaatagcct ctaaggtttt
     2821 aagttttata agaaaaaaaa gaatatataa ggcttttaaa gcttttaagg tttaacggtt
     2881 gtggacaaca agccagggat gtaacgcact gagaagccct tagagcctct caaagcaatt
     2941 ttgagtgaca caggaacact taacggctga catgggaatt agccatgggc ccgtgcgaat
     3001 catttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     3061 ccccggattc tgtggataac cgtattaccg cctggaccaa aacgaaaaaa caccctttcg
     3121 ggtgtctttt ctggaatttg gtaccgagcc tttggtcgaa aaaaaaagcc cgcactgtca
     3181 ggtgcgggct tttttctgtg tttccagcca agagcagtgc cttgcagaag ttctcgggcg
     3241 a
//
