LOCUS       sequence-423557-        3875 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  integration vector for three gene cassette with integration site E2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3875)
  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 3875)
  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: pCfB12714
FEATURES             Location/Qualifiers
     source          1..3875
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             476..487
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     rep_origin      complement(1274..1862)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1354..1373)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        1965..2069
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2070..2138,2139..2930)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2288..2307)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      complement(3505..3532)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(3548..3619)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(join(3631..3875,1..469))
                     /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 gtttatcggc ggtgatgtaa acattgtggc tgttaaaatt gtattccagc ttatggccca
       61 ggatattgcc gtcctcttta aagtcaatgc ctttcagctc aatgcggttt accagggtat
      121 cgccttcaaa tttcacttcc gcacgcgttt tgtacgtgcc gtcatcctta aaggaaatcg
      181 tgcgttcctg cacatagcct tccggcatgg cggacttgaa gaagtcatgc tgcttcatat
      241 ggtccggata acgagcaaag cactgaacac cataagtcag cgtcgttacc agagtcggcc
      301 aaggaaccgg cagtttacca gtagtacaga tgaacttcag cgtcagttta ccattagttg
      361 cgtcaccttc accctcgcca cgcacggaaa acttatgacc gttgacatca ccatccagtt
      421 ccaccagaat agggacgaca ccagtgaaca gctcttcgcc tttacgcatc tagtatttct
      481 cctctttctc tagtagctaa gatccgaacc tgccgtttct tgagttgccg cgatgttaag
      541 aaaacattca taaattaaat gtgaattgcc gcacacatta ttaaataaga tttacaaaat
      601 gttcaaaatg acgcatgaaa tcacgtttca ctttccgtct catcagtggt ctcacgttcc
      661 gtcttagtgc agcaaaatcg gacgatttct ccagagcatt cttgtcttat ggctatacag
      721 tccattccat gatgctgctt cctgcaatat gcgtagctgg caggtgtaga tttgccgaaa
      781 tcaagcggtc gcctattgaa actgtaagca accacccgga aggtgtttgt ggtgtacagt
      841 acttggagat gtataggcgt ggagagcacc agaatctatc ttggagacac cagacggagt
      901 tgttgaagtt ctcggacaaa agaaacacca gtaaagtcct ttgaacttct tgtggatcct
      961 attcaagctc cagtgaccct ctgatctgga taaaaatagc cctttcaaca tcaccacgga
     1021 actggtaggg gctggatgga ccaacactcc gtcctgagac atagatacaa agttgagagg
     1081 tcggacaaga aagttgacgg ccatggtatc gactgaatcg aatatcttca ttgtgtcaaa
     1141 gtgacaagcg atttggcacc atgcggccgc aggcacggtt atccacagaa tcaggggata
     1201 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     1261 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     1321 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     1381 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     1441 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     1501 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     1561 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     1621 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     1681 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     1741 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     1801 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     1861 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     1921 aagggatttt ggtcatgaca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     1981 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     2041 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     2101 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     2161 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     2221 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     2281 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     2341 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     2401 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     2461 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     2521 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     2581 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     2641 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     2701 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     2761 ataaatctgg agccggtgag cgtgggtccc gcggtatcat tgcagcactg gggccagatg
     2821 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     2881 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     2941 aagtttactc atatatactt tagatttatt taaaacttca tttttaattt aaaaggatct
     3001 aggtgaagat cctttttgat aatccgcagc ggccgcgaaa tacgccaggt ttgtagacta
     3061 taccaacagg agggttgtta acttctcggt agacacgtac ggcacccaca gaaggcatcc
     3121 cccctcccac tcctcgtcca gagacttcag actctcagtt cctcatcgct ggagttgctt
     3181 tttcagcgcg tttcttttgc aaatgtcgct ctttactctt ctggtccaaa cgggtaattg
     3241 cttggacggt caccatcgcg gttctccctg caccaaacac atgccaaaca catgccaaac
     3301 acatgtccag cgggctgact ggccgcatag ctaggcagcc aacagggtgc aaaaatgtaa
     3361 atagacagcc agggtcctaa atttagctgg tggtgagcgc tctctcctac aagtagtatc
     3421 ttatgctaac tgccgccgta tggccactct catttcttgc atctgtatgc caactccagc
     3481 tgagaccgaa ctgagacgta taaacgcaga aaggcccacc cgaaggtgag ccagtgtgac
     3541 tctagtagag agcgttcacc gacaaacaac agataaaacg aaaggcccag tctttcgact
     3601 gagcctttcg ttttatttga tgcctggtca tttgtacagt tcatccatac catgcgtgat
     3661 gcccgctgcg gttacgaact ccagcagaac catatgatcg cgtttctcgt tcggatcttt
     3721 agacagaacg ctttgcgtgc tcagatagtg attgtctggc agcagaacag gaccatcacc
     3781 gattggagtg ttttgctggt agtgatcagc cagctgcacg ctgccatcct ccacgttgtg
     3841 gcgaatttta aaattcgctt taatgccatt ttttt
//
