LOCUS       sequence-252449-        5973 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Z. rouxii centromeric vector containing G418 resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5973)
  AUTHORS   Bizzarri M, Cassanelli S, Duskova M, Sychrova H, Solieri L
  TITLE     A set of plasmids carrying antibiotic resistance markers and Cre 
            recombinase for genetic engineering of nonconventional yeast 
            Zygosaccharomyces rouxii.
  JOURNAL   Yeast. 2019 Dec;36(12):711-722. doi: 10.1002/yea.3438. Epub 2019 Sep
            11.
  PUBMED    31414502
REFERENCE   2  (bases 1 to 5973)
  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"; doi: "10.1002/yea.3438"; 
            journalName: "Yeast"; date: "2019-12"; volume: "36"; issue: "12"; 
            pages: "711-722"
COMMENT     Sequence Label: pZCAG
FEATURES             Location/Qualifiers
     source          1..5973
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    106..127
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(142..147,148..165,166..172)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    180..196
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     185..207
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     204..220
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             216..521
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIIAGPFS"
     misc_feature    233..289
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(290..307)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(290..306)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(299..321)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     RBS             1261..1269
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     protein_bind    2168..2201
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            2256..3612
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        2256..2599
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             2600..3409
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2667..2686)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      3415..3612
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    3675..3708
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        complement(3741..3956)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(3981..3999)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4067..4171
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4172..4240,4241..5032)
                     /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(4390..4409)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5203..5791
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5692..5711
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5945..5962
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc
      241 atgcctgcag gtcgactcta gaggatcccc gggtaccgag ctcgaattca ctggccgtcg
      301 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
      361 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
      421 agttgcgcag cctgaatggc gaatggcgcc tgatgcggta ttttctcctt acgcatctgt
      481 gcggtatttc acaccgcata atcgctgggc cattctcatg aagaatatct tgaatttatt
      541 gtcatattac tagtaggcat ttgtttatta ccagcttacg atataaagtc gttcatcttc
      601 tttatggcga gaaactgtaa ttgcgaaaat tgtctaggca atttttttca gtagttagtc
      661 tgagattttt ccttgtgaac caaccttcct gtcgttatta atactagaag ataagacgaa
      721 acagtttagt ttgtaggtac gtaaaaactt gttcaaagta ggcttcagta atcttgttcc
      781 gaaaaagttt ctcacatatc aaacaaatta gaggtttcgg aagtaagcca caacaaaaat
      841 aatgaatatt ttttaaacgc ttttaaaaca gataattatc aaaaaataga attaaagctt
      901 tatttgtatt aattttgaaa gtattaaata aaattatata ctgtcttata atatatatta
      961 ttatatcatg tgattaaatt acacagcttc taaagctctt taatggattg cggtttttac
     1021 atctactcta ctttagatca tactttgtct ttgattaaag tatattaagt ggccatctgt
     1081 gaccagtatc ttttgaaaca atttgtcata atatccatat tccatcatat taaacattta
     1141 cgttttgttc tactttaacc aatttgaaag acattattta aatcaaaaat attttctccg
     1201 agttagctat gaaaaagatt gaatacctca tgtataaaga tatcctctcc ttccaagaat
     1261 acctccttaa gtttcatttc taccgtcagg actcgtgttt tcgccaggag cgataagggg
     1321 acgtgtgatt tctgcatcat cgttattagg tgacactaag ttagtgcccg atcgatgcta
     1381 gcaagaatcg ggtcattgta gcgtatgcgc ctgtgaacat tctcttcaac aagtttgatt
     1441 ccattgcggt gaaatggtaa aagtcaaccc cctgcgatgt atattttcct gtacaatcaa
     1501 tcaaaaagcc aaatgattta gcattatctt tacatcttgt tattttacag attttatgtt
     1561 tagatctttt atgcttgctt ttcaaaaggc ttgcaggcaa gtgcacaaac aatacttaaa
     1621 taaatactac tcagtaataa cctatttctt agcatttttg acgaaatttg ctattttgtt
     1681 agagtctttt acaccatttg tctccacacc tccgcttaca tcaacaccaa taacgccatt
     1741 taatctaagc gcatcaccaa cattttctgg cgtcagtcca ccagctaaca taaaatgtaa
     1801 gctctcgggg ctctcttgcc ttccaaccca gtcagaaatc gagttccaat ccaaaagttc
     1861 acctgtccca cctgcttctg aatcaaacaa gggaataaac gaatgaggtt tctgtgaagc
     1921 tgcactgagt agtatgttgc agtcttttgg aaatacgagt cttttaataa ctggcaaacc
     1981 gaggaactct tggtattctt gccacgactc atctccatgc agttggacga tcgatgataa
     2041 gctgtcaaac atgagaattg ggtaataact gatataatta aattgaagct ctaatttgtg
     2101 agtttagtat acatgcattt acttataata cagttttttc gtacgctgca ggtcgacaac
     2161 ccttaatata acttcgtata atgtatgcta tacgaagtta ttaggtctag agatctgttt
     2221 agcttgcctc gtccccgccg ggtcacccgg ccagcgacat ggaggcccag aataccctcc
     2281 ttgacagtct tgacgtgcgc agctcagggg catgatgtga ctgtcgcccg tacatttagc
     2341 ccatacatcc ccatgtataa tcatttgcat ccatacattt tgatggccgc acggcgcgaa
     2401 gcaaaaatta cggctcctcg ctgcagacct gcgagcaggg aaacgctccc ctcacagacg
     2461 cgttgaattg tccccacgcc gcgcccctgt agagaaatat aaaaggttag gatttgccac
     2521 tgaggttctt ctttcatata cttcctttta aaatcttgct aggatacagt tctcacatca
     2581 catccgaaca taaacaacca tgggtaagga aaagactcac gtttcgaggc cgcgattaaa
     2641 ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc
     2701 aggtgcgaca atctatcgat tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca
     2761 tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac
     2821 ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt
     2881 actcaccact gcgatccccg gcaaaacagc attccaggta ttagaagaat atcctgattc
     2941 aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt
     3001 ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat
     3061 gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga
     3121 acaagtctgg aaagaaatgc ataagctttt gccattctca ccggattcag tcgtcactca
     3181 tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga
     3241 tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct
     3301 cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc
     3361 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaat cagtactgac
     3421 aataaaaaga ttcttgtttt caagaacttg tcatttgtat agttttttta tattgtagtt
     3481 gttctatttt aatcaaatgt tagcgtgatt tatatttttt ttcgcctcga catcatctgc
     3541 ccagatgcga agttaagtgc gcagaaagta atatcatgcg tcaatcgtat gtgaatgctg
     3601 gtcgctatac tgctgtcgat tcgatactaa cgccgccatc cagtgtcgaa aacgagctct
     3661 cgagaaccct taatataact tcgtataatg tatgctatac gaagttatta ggtgatatca
     3721 gatccactag tggcctatgc gatttatctt cgtttcctgc atgtttttgt tctgtgcagt
     3781 tgggttaaga atactgggca atttcatgtt tcttcaacac tacatatgcg tatatatacc
     3841 aatctaagtc tgtgctcctt ccttcgttct tccttctgtt cggagattac cgaatcaaaa
     3901 aaatttcaag gaaaccgaaa tcaaaaaaaa gaataaaaaa aaaatgatga attgaaaagc
     3961 taattcttga agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     4021 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     4081 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     4141 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     4201 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     4261 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     4321 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     4381 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     4441 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     4501 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     4561 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     4621 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     4681 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     4741 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     4801 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     4861 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     4921 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     4981 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     5041 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     5101 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     5161 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     5221 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     5281 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     5341 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     5401 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     5461 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     5521 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     5581 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     5641 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     5701 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     5761 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     5821 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     5881 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     5941 gcgcagcgag tcagtgagcg aggaagcgga aga
//
