LOCUS       sequence-317071-        4966 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low-copy expression vector for K. marxianus, G418 selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4966)
  AUTHORS   Rajkumar AS, Morrissey JP
  TITLE     Rational engineering of Kluyveromyces marxianus to create a chassis 
            for the production of aromatic products.
  JOURNAL   Microb Cell Fact. 2020 Nov 11;19(1):207. doi: 
            10.1186/s12934-020-01461-7.
  PUBMED    33176787
REFERENCE   2  (bases 1 to 4966)
  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: "Microb Cell
            Fact."; date: "2020-11-11"; volume: "19(1):207. doi"; pages: " 
            10.1186/s12934-020-01461-7"
COMMENT     Sequence Label: pMTU-DO-G418
FEATURES             Location/Qualifiers
     source          1..4966
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             347..358
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             365..1078
                     /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      1090..1161
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1177..1204
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     gene            1455..2810
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     CDS             1799..2608
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     terminator      2613..2810
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(3245..4060)
                     /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             complement(3245..4060)
                     /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     4052..4071
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4285..4873
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4774..4793
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aaccgtgcgg ccgcccctga attcgcatct agactgatga gacgtggtag agccacaaac
       61 agccggtaca agcaacgatc tccaggacca tctgaatcat gcgcggatga cacgaactca
      121 cgacggcgat cacagacatt aacccacagt acagacactg cgacaacgtg gcaattcgtc
      181 gcaatacaac gtgagaccga aagtgaaacg tgatttcatg cgtcattttg aacattttgt
      241 aaatcttatt taataatgtg tgcggcaatt cacatttaat ttatgaatgt tttcttaaca
      301 tcgcggcaac tcaagaaacg gcaggttcgg atcttagcta ctagagaaag aggagaaata
      361 ctagatgcgt aaaggcgaag agctgttcac tggtgtcgtc cctattctgg tggaactgga
      421 tggtgatgtc aacggtcata agttttccgt gcgtggcgag ggtgaaggtg acgcaactaa
      481 tggtaaactg acgctgaagt tcatctgtac tactggtaaa ctgccggttc cttggccgac
      541 tctggtaacg acgctgactt atggtgttca gtgctttgct cgttatccgg accatatgaa
      601 gcagcatgac ttcttcaagt ccgccatgcc ggaaggctat gtgcaggaac gcacgatttc
      661 ctttaaggat gacggcacgt acaaaacgcg tgcggaagtg aaatttgaag gcgataccct
      721 ggtaaaccgc attgagctga aaggcattga ctttaaagag gacggcaata tcctgggcca
      781 taagctggaa tacaatttta acagccacaa tgtttacatc accgccgata aacaaaaaaa
      841 tggcattaaa gcgaatttta aaattcgcca caacgtggag gatggcagcg tgcagctggc
      901 tgatcactac cagcaaaaca ctccaatcgg tgatggtcct gttctgctgc cagacaatca
      961 ctatctgagc acgcaaagcg ttctgtctaa agatccgaac gagaaacgcg atcatatggt
     1021 tctgctggag ttcgtaaccg cagcgggcat cacgcatggt atggatgaac tgtacaaatg
     1081 accaggcatc aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt
     1141 tgtttgtcgg tgaacgctct ctactagagt cacactggct caccttcggg tgggcctttc
     1201 tgcgtttata ggtctcagct ggaaatctgc tcgtcagtgg tgctcacact gacgaatcat
     1261 gtacagatca taccgatgac tgcctggcga ctcacaacta agcaagacag ccggaaccag
     1321 cgccggcgaa caccactgca tatatggcat atcacaacag tccacgtctc aagcagttac
     1381 agagatgtta cgaaccacta gtgcactgca gtacagttta gcttgcctcg tccccgccgg
     1441 gtcacccggc cagcgacatg gaggcccaga ataccctcct tgacagtctt gacgtgcgca
     1501 gctcaggggc atgatgtgac tgtcgcccgt acatttagcc catacatccc catgtataat
     1561 catttgcatc catacatttt gatggccgca cggcgcgaag caaaaattac ggctcctcgc
     1621 tccagacctg cgagcaggga aacgctcccc tcacagacgc gttgaattgt ccccacgccg
     1681 cgcccctgta gagaaatata aaaggttagg atttgccact gaggttcttc tttcatatac
     1741 ttccttttaa aatcttgcta ggatacagtt ctcacatcac atccgaacat aaacaaaaat
     1801 gggtaaggaa aagactcacg tttcgaggcc gcgattaaat tccaacatgg atgctgattt
     1861 atatgggtat aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt
     1921 gtatgggaag cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa
     1981 tgatgttaca gatgagatgg tcagactaaa ctggctgacg gaatttatgc ctcttccgac
     2041 catcaagcat tttatccgta ctcctgatga tgcatggtta ctcaccactg cgatccccgg
     2101 caaaacagca ttccaggtat tagaagaata tcctgattca ggtgaaaata ttgttgatgc
     2161 gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc cttttaacag
     2221 cgatcgcgta tttcgtctgg ctcaggcgca atcacgaatg aataacggtt tggttgatgc
     2281 gagtgatttt gatgacgagc gtaatggctg gcctgttgaa caagtctgga aagaaatgca
     2341 taagcttttg ccattctcac cggattcagt cgtcactcat ggtgatttct cacttgataa
     2401 ccttattttt gacgagggga aattaatagg ttgtattgat gttggacgag tcggaatcgc
     2461 agaccgatac caggatcttg ccatcctatg gaactgcctc ggtgagtttt ctccttcatt
     2521 acagaaacgg ctttttcaaa aatatggtat tgataatcct gatatgaata aattgcagtt
     2581 tcatttgatg ctcgatgagt ttttctaaag taactgacaa taaaaagatt cttgttttca
     2641 agaacttgtc atttgtatag tttttttata ttgtagttgt tctattttaa tcaaatgtta
     2701 gcgtgattta tatttttttt cgcctcgaca tcatctgccc agatgcgaag ttaagtgcgc
     2761 agaaagtaat atcatgcgtc aatcgtatgt gaatgctggt cgctatactg gagtttctac
     2821 gcaagacttc ttgaagtgaa aaccaacttt cagtcttcaa actaaaaatg aaaatcagtg
     2881 ggctaaattc agcataatac ctgtggatat aattacatac aaaaatataa ttaaaaatat
     2941 acgttaaaaa taatttattt ttgtaaaaac tataaaatat attttacttt cggaacaact
     3001 ttttaactta taattttgtt ttaaataaaa acattgtatt taaaaataat aaattattaa
     3061 ataaaatttt aaagtatgtt tatttaaaat atagaatact attaaaatta ctttaaactt
     3121 taaaataaaa aaataataaa ttatatagtt ttaaaattaa taaattttat acacgtgacc
     3181 agatcataat agttttcttt ccttgccgag cggccgcgtg ttacaaccaa ttaaccaatt
     3241 ctgattagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat
     3301 caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt
     3361 tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac
     3421 aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga
     3481 cgactgaatc cggtgagaat ggcaaaagct tatgcatttc tttccagact tgttcaacag
     3541 gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg
     3601 attgcgcctg agcgaggcga aatacgcgat cgctgttaaa aggacaatta caaacaggaa
     3661 tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag
     3721 gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg
     3781 catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc
     3841 agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca
     3901 gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc
     3961 cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc
     4021 gcggcctgga gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt
     4081 ttatgtaagc agacagtttt attgttcatg atgatatatt tttatcttgt gcaatgtaac
     4141 atcagagatt ttgagacaca acgtggcttt gttgaataaa tcgaactttt gctgagttga
     4201 aggatcagtc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     4261 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     4321 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     4381 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     4441 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     4501 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     4561 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     4621 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     4681 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     4741 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     4801 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     4861 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     4921 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggat
//
