LOCUS       sequence-424071-        3960 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  integration vector for single cassette with integration site E2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3960)
  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 3960)
  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: pCfB12713
FEATURES             Location/Qualifiers
     source          1..3960
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             401..1192
                     /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(1700..1727)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(1743..1814)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1826..2539)
                     /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"
     RBS             2546..2557
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     rep_origin      complement(3343..3931)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3423..3442)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgacca gccaggacag
       61 aaatgcctcg acttcgctgc tacccaaggt tgccgggtga cgcacaccgt ggaaacggat
      121 gaaggcacga acccagtgga cataagcctg ttcggttcgt aagctgtaat gcaagtagcg
      181 tatgcgctca cgcaactggt ccagaacctt gaccgaacgc agcggtggta acggcgcagt
      241 ggcggttttc atggcttgtt atgactgttt ttttggggta cagtctatgc ctcgggcatc
      301 caagcagcaa gcgcgttacg ccgtgggtcg atgtttgatg ttatggagca gcaacgatgt
      361 tacgcagcag ggcagtcgcc ctaaaacaaa gttaaacatt atgagggaag cggtgatcgc
      421 cgaagtatcg actcaactat cagaggtagt tggcgtcatc gagcgccatc tcgaaccgac
      481 gttgctggcc gtacatttgt acggctccgc agtggatggc ggcctgaagc cacacagtga
      541 tattgatttg ctggttacgg tgaccgtaag gcttgatgaa acaacgcggc gagctttgat
      601 caacgacctt ttggaaactt cggcttcccc tggagagagc gagattctcc gcgctgtaga
      661 agtcaccatt gttgtgcacg acgacatcat tccgtggcgt tatccagcta agcgcgaact
      721 gcaatttgga gaatggcagc gcaatgacat tcttgcgggt atcttcgagc cagccacgat
      781 cgacattgat ctggctatct tgctgacaaa agcaagagaa catagcgttg ccttggtagg
      841 tccagcggcg gaggaactct ttgatccggt tcctgaacag gatctatttg aggcgctaaa
      901 tgaaacctta acgctatgga actcgccgcc cgactgggct ggcgatgagc gaaatgtagt
      961 gcttacgttg tcccgcattt ggtacagcgc agtaaccggc aaaatcgcgc cgaaggatgt
     1021 cgctgccgac tgggcaatgg agcgcctgcc ggcccagtat cagcccgtca tacttgaagc
     1081 tagacaggct tatcttggac aagaagaaga tcgcttggcc tcgcgcgcag atcagttgga
     1141 agaatttgtc cactacgtga aaggcgagat caccaaggta gtcggcaaat aatcagaatt
     1201 ggttaattgg ttgtaacacc gcagcggccg cgaaatacgc caggtttgta gactatacca
     1261 acaggagggt tgttaacttc tcggtagaca cgtacggcac ccacagaagg catcccccct
     1321 cccactcctc gtccagagac ttcagactct cagttcctca tcgctggagt tgctttttca
     1381 gcgcgtttct tttgcaaatg tcgctcttta ctcttctggt ccaaacgggt aattgcttgg
     1441 acggtcacca tcgcggttct ccctgcacca aacacatgcc aaacacatgc caaacacatg
     1501 tccagcgggc tgactggccg catagctagg cagccaacag ggtgcaaaaa tgtaaataga
     1561 cagccagggt cctaaattta gctggtggtg agcgctctct cctacaagta gtatcttatg
     1621 ctaactgccg ccgtatggcc actctcattt cttgcatctg tatgccaact ccgtctccga
     1681 acactctgag acctataaac gcagaaaggc ccacccgaag gtgagccagt gtgactctag
     1741 tagagagcgt tcaccgacaa acaacagata aaacgaaagg cccagtcttt cgactgagcc
     1801 tttcgtttta tttgatgcct ggtcatttgt acagttcatc cataccatgc gtgatgcccg
     1861 ctgcggttac gaactccagc agaaccatat gatcgcgttt ctcgttcgga tctttagaca
     1921 gaacgctttg cgtgctcaga tagtgattgt ctggcagcag aacaggacca tcaccgattg
     1981 gagtgttttg ctggtagtga tcagccagct gcacgctgcc atcctccacg ttgtggcgaa
     2041 ttttaaaatt cgctttaatg ccattttttt gtttatcggc ggtgatgtaa acattgtggc
     2101 tgttaaaatt gtattccagc ttatggccca ggatattgcc gtcctcttta aagtcaatgc
     2161 ctttcagctc aatgcggttt accagggtat cgccttcaaa tttcacttcc gcacgcgttt
     2221 tgtacgtgcc gtcatcctta aaggaaatcg tgcgttcctg cacatagcct tccggcatgg
     2281 cggacttgaa gaagtcatgc tgcttcatat ggtccggata acgagcaaag cactgaacac
     2341 cataagtcag cgtcgttacc agagtcggcc aaggaaccgg cagtttacca gtagtacaga
     2401 tgaacttcag cgtcagttta ccattagttg cgtcaccttc accctcgcca cgcacggaaa
     2461 acttatgacc gttgacatca ccatccagtt ccaccagaat agggacgaca ccagtgaaca
     2521 gctcttcgcc tttacgcatc tagtatttct cctctttctc tagtagctaa gatccgaacc
     2581 tgccgtttct tgagttgccg cgatgttaag aaaacattca taaattaaat gtgaattgcc
     2641 gcacacatta ttaaataaga tttacaaaat gttcaaaatg acgcatgaaa tcacgtttca
     2701 ctttcggtct ccccgtcatc cgagacgccg tcttagtgca gcaaaatcgg acgatttctc
     2761 cagagcattc ttgtcttatg gctatacagt ccattccatg atgctgcttc ctgcaatatg
     2821 cgtagctggc aggtgtagat ttgccgaaat caagcggtcg cctattgaaa ctgtaagcaa
     2881 ccacccggaa ggtgtttgtg gtgtacagta cttggagatg tataggcgtg gagagcacca
     2941 gaatctatct tggagacacc agacggagtt gttgaagttc tcggacaaaa gaaacaccag
     3001 taaagtcctt tgaacttctt gtggatccta ttcaagctcc agtgaccctc tgatctggat
     3061 aaaaatagcc ctttcaacat caccacggaa ctggtagggg ctggatggac caacactccg
     3121 tcctgagaca tagatacaaa gttgagaggt cggacaagaa agttgacggc catggtatcg
     3181 actgaatcga atatcttcat tgtgtcaaag tgacaagcga tttggcacca tgcggccgca
     3241 ggcacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     3301 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     3361 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     3421 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     3481 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     3541 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     3601 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     3661 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     3721 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     3781 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     3841 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     3901 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
//
