LOCUS       sequence-424243-        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: pCfB12712
FEATURES             Location/Qualifiers
     source          1..3960
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(72..863)
                     /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"
     rep_origin      1293..1881
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1782..1801
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     RBS             2667..2678
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2685..3398
                     /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"
     terminator      3410..3481
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3497..3524
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
ORIGIN
        1 tgttggtata gtctacaaac ctggcgtatt tcgcggccgc tgcggtgtta caaccaatta
       61 accaattctg attatttgcc gactaccttg gtgatctcgc ctttcacgta gtggacaaat
      121 tcttccaact gatctgcgcg cgaggccaag cgatcttctt cttgtccaag ataagcctgt
      181 ctagcttcaa gtatgacggg ctgatactgg gccggcaggc gctccattgc ccagtcggca
      241 gcgacatcct tcggcgcgat tttgccggtt actgcgctgt accaaatgcg ggacaacgta
      301 agcactacat ttcgctcatc gccagcccag tcgggcggcg agttccatag cgttaaggtt
      361 tcatttagcg cctcaaatag atcctgttca ggaaccggat caaagagttc ctccgccgct
      421 ggacctacca aggcaacgct atgttctctt gcttttgtca gcaagatagc cagatcaatg
      481 tcgatcgtgg ctggctcgaa gatacccgca agaatgtcat tgcgctgcca ttctccaaat
      541 tgcagttcgc gcttagctgg ataacgccac ggaatgatgt cgtcgtgcac aacaatggtg
      601 acttctacag cgcggagaat ctcgctctct ccaggggaag ccgaagtttc caaaaggtcg
      661 ttgatcaaag ctcgccgcgt tgtttcatca agccttacgg tcaccgtaac cagcaaatca
      721 atatcactgt gtggcttcag gccgccatcc actgcggagc cgtacaaatg tacggccagc
      781 aacgtcggtt cgagatggcg ctcgatgacg ccaactacct ctgatagttg agtcgatact
      841 tcggcgatca ccgcttccct cataatgttt aactttgttt tagggcgact gccctgctgc
      901 gtaacatcgt tgctgctcca taacatcaaa catcgaccca cggcgtaacg cgcttgctgc
      961 ttggatgccc gaggcataga ctgtacccca aaaaaacagt cataacaagc catgaaaacc
     1021 gccactgcgc cgttaccacc gctgcgttcg gtcaaggttc tggaccagtt gcgtgagcgc
     1081 atacgctact tgcattacag cttacgaacc gaacaggctt atgtccactg ggttcgtgcc
     1141 ttcatccgtt tccacggtgt gcgtcacccg gcaaccttgg gtagcagcga agtcgaggca
     1201 tttctgtcct ggctggtcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     1261 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     1321 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     1381 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     1441 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     1501 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     1561 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     1621 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     1681 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     1741 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     1801 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     1861 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     1921 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     1981 gcctgcggcc gcatggtgcc aaatcgcttg tcactttgac acaatgaaga tattcgattc
     2041 agtcgatacc atggccgtca actttcttgt ccgacctctc aactttgtat ctatgtctca
     2101 ggacggagtg ttggtccatc cagcccctac cagttccgtg gtgatgttga aagggctatt
     2161 tttatccaga tcagagggtc actggagctt gaataggatc cacaagaagt tcaaaggact
     2221 ttactggtgt ttcttttgtc cgagaacttc aacaactccg tctggtgtct ccaagataga
     2281 ttctggtgct ctccacgcct atacatctcc aagtactgta caccacaaac accttccggg
     2341 tggttgctta cagtttcaat aggcgaccgc ttgatttcgg caaatctaca cctgccagct
     2401 acgcatattg caggaagcag catcatggaa tggactgtat agccataaga caagaatgct
     2461 ctggagaaat cgtccgattt tgctgcacta agacggcgtc tcgccaaacg gggagaccga
     2521 aagtgaaacg tgatttcatg cgtcattttg aacattttgt aaatcttatt taataatgtg
     2581 tgcggcaatt cacatttaat ttatgaatgt tttcttaaca tcgcggcaac tcaagaaacg
     2641 gcaggttcgg atcttagcta ctagagaaag aggagaaata ctagatgcgt aaaggcgaag
     2701 agctgttcac tggtgtcgtc cctattctgg tggaactgga tggtgatgtc aacggtcata
     2761 agttttccgt gcgtggcgag ggtgaaggtg acgcaactaa tggtaaactg acgctgaagt
     2821 tcatctgtac tactggtaaa ctgccggttc cttggccgac tctggtaacg acgctgactt
     2881 atggtgttca gtgctttgct cgttatccgg accatatgaa gcagcatgac ttcttcaagt
     2941 ccgccatgcc ggaaggctat gtgcaggaac gcacgatttc ctttaaggat gacggcacgt
     3001 acaaaacgcg tgcggaagtg aaatttgaag gcgataccct ggtaaaccgc attgagctga
     3061 aaggcattga ctttaaagag gacggcaata tcctgggcca taagctggaa tacaatttta
     3121 acagccacaa tgtttacatc accgccgata aacaaaaaaa tggcattaaa gcgaatttta
     3181 aaattcgcca caacgtggag gatggcagcg tgcagctggc tgatcactac cagcaaaaca
     3241 ctccaatcgg tgatggtcct gttctgctgc cagacaatca ctatctgagc acgcaaagcg
     3301 ttctgtctaa agatccgaac gagaaacgcg atcatatggt tctgctggag ttcgtaaccg
     3361 cagcgggcat cacgcatggt atggatgaac tgtacaaatg accaggcatc aaataaaacg
     3421 aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct
     3481 ctactagagt cacactggct caccttcggg tgggcctttc tgcgtttata ggtctcagag
     3541 tgatgggaga cggagttggc atacagatgc aagaaatgag agtggccata cggcggcagt
     3601 tagcataaga tactacttgt aggagagagc gctcaccacc agctaaattt aggaccctgg
     3661 ctgtctattt acatttttgc accctgttgg ctgcctagct atgcggccag tcagcccgct
     3721 ggacatgtgt ttggcatgtg tttggcatgt gtttggtgca gggagaaccg cgatggtgac
     3781 cgtccaagca attacccgtt tggaccagaa gagtaaagag cgacatttgc aaaagaaacg
     3841 cgctgaaaaa gcaactccag cgatgaggaa ctgagagtct gaagtctctg gacgaggagt
     3901 gggagggggg atgccttctg tgggtgccgt acgtgtctac cgagaagtta acaaccctcc
//
