LOCUS       sequence-424074-        4016 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  integration vector for single cassette with integration site E3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4016)
  AUTHORS   Jiang W, Wang S, Avila P, Jorgensen TS, Yang Z, Borodina I
  TITLE     Combinatorial iterative method for metabolic engineering of Yarrowia
            lipolytica: application for betanin biosynthesis.
  JOURNAL   Metab Eng. 2024 Sep 9:S1096-7176(24)00110-1. doi: 
            10.1016/j.ymben.2024.09.003.
  PUBMED    39260817
REFERENCE   2  (bases 1 to 4016)
  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: "Metab Eng. 
            2024 Sep 9:S1096-7176(24)00110-1. doi: 10.1016/j.ymben.2024.09.003."
COMMENT     Sequence Label: pCfB13015
FEATURES             Location/Qualifiers
     source          1..4016
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             524..535
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     rep_origin      complement(1344..1932)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1424..1443)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             2362..3153
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     terminator      complement(3694..3721)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(3737..3808)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(join(3820..4016,1..517))
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
ORIGIN
        1 cctccacgtt gtggcgaatt ttaaaattcg ctttaatgcc atttttttgt ttatcggcgg
       61 tgatgtaaac attgtggctg ttaaaattgt attccagctt atggcccagg atattgccgt
      121 cctctttaaa gtcaatgcct ttcagctcaa tgcggtttac cagggtatcg ccttcaaatt
      181 tcacttccgc acgcgttttg tacgtgccgt catccttaaa ggaaatcgtg cgttcctgca
      241 catagccttc cggcatggcg gacttgaaga agtcatgctg cttcatatgg tccggataac
      301 gagcaaagca ctgaacacca taagtcagcg tcgttaccag agtcggccaa ggaaccggca
      361 gtttaccagt agtacagatg aacttcagcg tcagtttacc attagttgcg tcaccttcac
      421 cctcgccacg cacggaaaac ttatgaccgt tgacatcacc atccagttcc accagaatag
      481 ggacgacacc agtgaacagc tcttcgcctt tacgcatcta gtatttctcc tctttctcta
      541 gtagctaaga tccgaacctg ccgtttcttg agttgccgcg atgttaagaa aacattcata
      601 aattaaatgt gaattgccgc acacattatt aaataagatt tacaaaatgt tcaaaatgac
      661 gcatgaaatc acgtttcact ttcggtctcc ccgtttggcg agacgccgtt gtttgatgtc
      721 ttgagtttga ggtcatttag actaggagta caagttcgta cgatacaggg aagaagtacc
      781 ggtgtattgt actggtactg tgagtacaca atcggcatga acgctcttcc aaacttcctg
      841 cgtcggtgta ccgtttgtgc gttctgtatt ggtgtcacaa tgtggcccat acttgtacaa
      901 gttgcatggt atatagtgcg agctgtttcg tataatgtgg tacatacaca tgcactgaag
      961 gtacctgaac gtatacgtcg cattgaaaac gggtctgtat gttgtattgt gcgaacatgg
     1021 caacataatg tagtgacgat acttgtaacg tcactgctag aacatttctc ctcagtaaac
     1081 catggataga agccgcttat aacagtatcg atagcacaat tgtcactggt gccaacgtac
     1141 tgtgtatgta ccgcacagta ccagtacgag cgagtatcag tacttacgta tacgacacct
     1201 gaccgtggag aacagagtca cgaaaaaagt tagcggccgc aggcacggtt atccacagaa
     1261 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     1321 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     1381 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     1441 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     1501 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     1561 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     1621 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     1681 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     1741 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     1801 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     1861 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     1921 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     1981 aactcacgtt aagggatttt ggtcatgacc agccaggaca gaaatgcctc gacttcgctg
     2041 ctacccaagg ttgccgggtg acgcacaccg tggaaacgga tgaaggcacg aacccagtgg
     2101 acataagcct gttcggttcg taagctgtaa tgcaagtagc gtatgcgctc acgcaactgg
     2161 tccagaacct tgaccgaacg cagcggtggt aacggcgcag tggcggtttt catggcttgt
     2221 tatgactgtt tttttggggt acagtctatg cctcgggcat ccaagcagca agcgcgttac
     2281 gccgtgggtc gatgtttgat gttatggagc agcaacgatg ttacgcagca gggcagtcgc
     2341 cctaaaacaa agttaaacat tatgagggaa gcggtgatcg ccgaagtatc gactcaacta
     2401 tcagaggtag ttggcgtcat cgagcgccat ctcgaaccga cgttgctggc cgtacatttg
     2461 tacggctccg cagtggatgg cggcctgaag ccacacagtg atattgattt gctggttacg
     2521 gtgaccgtaa ggcttgatga aacaacgcgg cgagctttga tcaacgacct tttggaaact
     2581 tcggcttccc ctggagagag cgagattctc cgcgctgtag aagtcaccat tgttgtgcac
     2641 gacgacatca ttccgtggcg ttatccagct aagcgcgaac tgcaatttgg agaatggcag
     2701 cgcaatgaca ttcttgcggg tatcttcgag ccagccacga tcgacattga tctggctatc
     2761 ttgctgacaa aagcaagaga acatagcgtt gccttggtag gtccagcggc ggaggaactc
     2821 tttgatccgg ttcctgaaca ggatctattt gaggcgctaa atgaaacctt aacgctatgg
     2881 aactcgccgc ccgactgggc tggcgatgag cgaaatgtag tgcttacgtt gtcccgcatt
     2941 tggtacagcg cagtaaccgg caaaatcgcg ccgaaggatg tcgctgccga ctgggcaatg
     3001 gagcgcctgc cggcccagta tcagcccgtc atacttgaag ctagacaggc ttatcttgga
     3061 caagaagaag atcgcttggc ctcgcgcgca gatcagttgg aagaatttgt ccactacgtg
     3121 aaaggcgaga tcaccaaggt agtcggcaaa taatcagaat tggttaattg gttgtaacac
     3181 cgcagcggcc gcgtgaagga aatgcctaaa acctgaattg aacgaagcgc ttttctacaa
     3241 gtaccgtacg tgtaaggtgg atggtgggcc tgatgtgtta gtttgactgt tctactacgt
     3301 gtcaccttac tgtgcctgta ctgtacttga agatggagca atatgtacac atgcgcgaaa
     3361 cttcaggagt caggttgtta tgcaacagga aatttttatg gtgatgaatt ataattatgc
     3421 acaggtccca aatgggaaaa atgtatggag agaaaccttc gattggtgtt tgtggcggtg
     3481 caatctgggg aacttcgcgt gtagtatttg tacttgtagt agaagttctc catcatttta
     3541 caagaataac cattagtcaa tgggtagttg tatgaaatag aaccatcttt aaatgcattt
     3601 ccatcgctga tgaagaaaat ggcttagtta actgggtatg tgtggtcatt tctgtgccag
     3661 aactccgtct cccatcactc tgagacctat aaacgcagaa aggcccaccc gaaggtgagc
     3721 cagtgtgact ctagtagaga gcgttcaccg acaaacaaca gataaaacga aaggcccagt
     3781 ctttcgactg agcctttcgt tttatttgat gcctggtcat ttgtacagtt catccatacc
     3841 atgcgtgatg cccgctgcgg ttacgaactc cagcagaacc atatgatcgc gtttctcgtt
     3901 cggatcttta gacagaacgc tttgcgtgct cagatagtga ttgtctggca gcagaacagg
     3961 accatcaccg attggagtgt tttgctggta gtgatcagcc agctgcacgc tgccat
//
