LOCUS       sequence-147688-        3374 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Supplemental Figure 3. Integration Site 1..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3374)
  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 3374)
  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: PP74
FEATURES             Location/Qualifiers
     source          1..3374
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      414..661
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(430..448)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     rep_origin      complement(736..1324)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(816..835)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(1415..1448)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     protein_bind    2113..2160
                     /label=Bxb1 attP
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attP site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     promoter        2476..2889
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        2896..2943
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             join(2968..3374,1..409)
                     /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             join(2968..3374,1..409)
                     /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(3041..3060)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat
       61 gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga
      121 acaagtctgg aaagaaatgc ataaactttt gccattctca ccggattcag tcgtcactca
      181 tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga
      241 tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct
      301 cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc
      361 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaac acatcatgta
      421 attagttatg tcacgcttac attcacgccc tccccccaca tccgctctaa ccgaaaagga
      481 aggagttaga caacctgaag tctaggtccc tatttatttt tttatagtta tgttagtatt
      541 aagaacgtta tttatatttc aaatttttct tttttttctg tacagacgcg tgtacgcatg
      601 taacattata ctgaaaacct tgcttgagaa ggttttggga cgctcgaagg ctttaatttg
      661 caagctggag accaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
      721 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
      781 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
      841 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
      901 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
      961 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     1021 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     1081 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     1141 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     1201 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     1261 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     1321 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     1381 ttaagggatt ttggtcatgc atgagatcag atctgaagtt cctatacttt ctagagaata
     1441 ggaacttcaa gcttgtggaa cattgagacc aaacaagact cgcttcgatg ctttcagatc
     1501 cattttccca gcaggtaccg tctccggtgc tccgaaggta agagcaatgc aactcatagg
     1561 agaattggaa ggagaaaaga gaggtgttta tgcgggggcc gtaggacact ggtcgtacga
     1621 tggaaaatcg atggacacat gtattgcctt aagaacaatg gtcgtcaagg acggtgtcgc
     1681 ttaccttcaa gccggaggtg gaattgtcta cgattctgac ccctatgacg agtacatcga
     1741 aaccatgaac aaaatgagat ccaacaataa caccatcttg gaggctgaga aaatctggac
     1801 cgataggttg gccagagacg agaatcaaag tgaatccgaa gaaaacgatc aatgaacgga
     1861 ggacgtaagt aggaatttat gtaatcatgc caatacatct ttagatttct tcctcttctt
     1921 tttcatgaga ttattggaaa ccaccagaat cgaatataaa aggcgaacac ctttcccaat
     1981 tttggtttct cctgacccaa agactttaaa tttaatttat ttgtccctat ttcaatcaat
     2041 tgaacaacta tttcggctgg acggcgacgt aaacggccac aagtttatgg ccgtgatgac
     2101 ctgtgtcttc gtggtttgtc tggtcaacca ccgcggtctc agtggtgtac ggtacaaacc
     2161 caaagcagca cgacacggca actacaagac ccgcgccgag ggcatgttcc ccaaagcgat
     2221 accacttgaa gcagtggtac tgcttgtggg tacactctgc gggtgtgaag ttcgagggcg
     2281 acaccctggt gaaccgcatc gagctgaagg gcatcttcca actcgcttaa ttgcgagttt
     2341 ttatttcgtt tatttcaatt aaggtaacta aaaaactcct tttacacatg aagcagcacg
     2401 acttcttcaa gtccgccatg cccgaaaaac gcctcttcag agtacagaag attaagtgag
     2461 accttcgttt gtgcggatcc cccacacacc atagcttcaa aatgtttcta ctcctttttt
     2521 actcttccag attttctcgg actccgcgca tcgccgtacc acttcaaaac acccaagcac
     2581 agcatactaa atttcccctc tttcttcctc tagggtgtcg ttaattaccc gtactaaagg
     2641 tttggaaaag aaaaaagaga ccgcctcgtt tctttttctt cgtcgaaaaa ggcaataaaa
     2701 atttttatca cgtttctttt tcttgaaaat tttttttttt gatttttttc tctttcgatg
     2761 acctcccatt gatatttaag ttaataaacg gtcttcaatt tctcaagttt cagtttcatt
     2821 tttcttgttc tattacaact ttttttactt cttgctcatt agaaagaaag catagcaatc
     2881 taatctaagg gcggtgttga caattaatca tcggcatagt atatcggcat agtataatac
     2941 gacaaggtga ggaactaaac catggtaatg agccatattc aacgggaaac gtcttgctct
     3001 aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg ggctcgcgat
     3061 aatgtcgggc aatcaggtgc gacaatctat cgattgtatg ggaagcccga tgcgccagag
     3121 ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga gatggtcaga
     3181 ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat ccgtactcct
     3241 gatgatgcat ggttactcac cactgcgatc cccgggaaaa cagcattcca ggtattagaa
     3301 gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct gcgccggttg
     3361 cattcgattc ctgt
//
