LOCUS       sequence-171564-        3547 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Supplemental Figure 3. Integration Site 2..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3547)
  AUTHORS   Perez-Pinera P, Han N, Cleto S, Cao J, Purcell O, Shah KA, Lee K, 
            Ram R, Lu TK
  TITLE     Synthetic biology and microbioreactor platforms for programmable 
            production of biologics at the point-of-care.
  JOURNAL   Nat Commun. 2016 Jul 29;7:12211. doi: 10.1038/ncomms12211.
  PUBMED    27470089
REFERENCE   2  (bases 1 to 3547)
  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 
            Commun."; date: "2016-07-29"; volume: "7:12211. doi"; pages: " 
            10.1038/ncomms12211"
COMMENT     Sequence Label: PP75
FEATURES             Location/Qualifiers
     source          1..3547
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(304..892)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(384..403)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    1854..1901
                     /label=Bxb1 attP
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attP site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     promoter        2217..2630
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        2637..2684
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2709..3524
                     /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             2709..3524
                     /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(2782..2801)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      join(3529..3547,1..229)
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(join(3545..3547,1..16))
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
ORIGIN
        1 acgcttacat tcacgccctc cccccacatc cgctctaacc gaaaaggaag gagttagaca
       61 acctgaagtc taggtcccta tttatttttt tatagttatg ttagtattaa gaacgttatt
      121 tatatttcaa atttttcttt tttttctgta cagacgcgtg tacgcatgta acattatact
      181 gaaaaccttg cttgagaagg ttttgggacg ctcgaaggct ttaatttgca agctggagac
      241 caacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
      301 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
      361 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
      421 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
      481 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
      541 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
      601 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
      661 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
      721 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
      781 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
      841 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
      901 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
      961 ggtcatgcat gagatcagat cttgatacta cagataccat tgaatgtggg tgaaaagagg
     1021 tatcaaaaga attgcgtctt tcctcaaaat tatttatttc agaagtgtaa acccatgccg
     1081 aattgagagt tgtctgaaaa catcaatttt agaaccttct cctcgattga tttgtgtaga
     1141 ggttggcagt aacaggaaaa aatttatgga catagctttt ctttaccttt gaatgttggg
     1201 ttatccaaaa gagttccaga agacgcaatc cataagaaaa cagccaaaga acccaagtta
     1261 aaagctgata ctagctagca tgaaaacaat tcgttgcatt attgctcagg tttttaaaca
     1321 caaacagtca tgctaacaat cgaactcagt tcaaagttgt tggaaactgc tgtcgacggt
     1381 atataagtta tttcttgata aattgcaaag caagttagca cccatgaaaa actattacat
     1441 gtgattattt cgttattgac cttgacccaa ttagcagcat gaagcctgac gtaccagttc
     1501 ctgcacagta gtagctggtg tcaggtggga acttgttaat cctccagcat tcattcattt
     1561 gaaaaaacta aactcaaact tagccaggca cttgctgtca ccagagtgca aagcacactc
     1621 tacccagttt tataccttcc tggcgttttg ttattgtgaa cattcatgag attattggaa
     1681 accaccagaa tcgaatataa aaggcgaaca cctttcccaa ttttggtttc tcctgaccca
     1741 aagactttaa atttaattta tttgtcccta tttcaatcaa ttgaacaact atttcggctg
     1801 gacggcgacg taaacggcca caagtttatg gccgtgatga cctgtgtctt cgtggtttgt
     1861 ctggtcaacc accgcggtct cagtggtgta cggtacaaac ccaaagcagc acgacacggc
     1921 aactacaaga cccgcgccga gggcatgttc cccaaagcga taccacttga agcagtggta
     1981 ctgcttgtgg gtacactctg cgggtgtgaa gttcgagggc gacaccctgg tgaaccgcat
     2041 cgagctgaag ggcatcttcc aactcgctta attgcgagtt tttatttcgt ttatttcaat
     2101 taaggtaact aaaaaactcc ttttacacat gaagcagcac gacttcttca agtccgccat
     2161 gcccgaaaaa cgcctcttca gagtacagaa gattaagtga gaccttcgtt tgtgcggatc
     2221 ccccacacac catagcttca aaatgtttct actccttttt tactcttcca gattttctcg
     2281 gactccgcgc atcgccgtac cacttcaaaa cacccaagca cagcatacta aatttcccct
     2341 ctttcttcct ctagggtgtc gttaattacc cgtactaaag gtttggaaaa gaaaaaagag
     2401 accgcctcgt ttctttttct tcgtcgaaaa aggcaataaa aatttttatc acgtttcttt
     2461 ttcttgaaaa tttttttttt tgattttttt ctctttcgat gacctcccat tgatatttaa
     2521 gttaataaac ggtcttcaat ttctcaagtt tcagtttcat ttttcttgtt ctattacaac
     2581 tttttttact tcttgctcat tagaaagaaa gcatagcaat ctaatctaag ggcggtgttg
     2641 acaattaatc atcggcatag tatatcggca tagtataata cgacaaggtg aggaactaaa
     2701 ccatggtaat gagccatatt caacgggaaa cgtcttgctc taggccgcga ttaaattcca
     2761 acatggatgc tgatttatat gggtataaat gggctcgcga taatgtcggg caatcaggtg
     2821 cgacaatcta tcgattgtat gggaagcccg atgcgccaga gttgtttctg aaacatggca
     2881 aaggtagcgt tgccaatgat gttacagatg agatggtcag actaaactgg ctgacggaat
     2941 ttatgcctct tccgaccatc aagcatttta tccgtactcc tgatgatgca tggttactca
     3001 ccactgcgat ccccgggaaa acagcattcc aggtattaga agaatatcct gattcaggtg
     3061 aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta
     3121 attgtccttt taacagcgat cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata
     3181 acggtttggt tgatgcgagt gattttgatg acgagcgtaa tggctggcct gttgaacaag
     3241 tctggaaaga aatgcataaa cttttgccat tctcaccgga ttcagtcgtc actcatggtg
     3301 atttctcact tgataacctt atttttgacg aggggaaatt aataggttgt attgatgttg
     3361 gacgagtcgg aatcgcagac cgataccagg atcttgccat cctatggaac tgcctcggtg
     3421 agttttctcc ttcattacag aaacggcttt ttcaaaaata tggtattgat aatcctgata
     3481 tgaataaatt gcagtttcat ttgatgctcg atgagttttt ctaacacatc atgtaattag
     3541 ttatgtc
//
