LOCUS       sequence-66302-e        6645 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TEF1 and GAL10 promoter controlled expression cassettes with G418 
            resistance for yeast transformation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6645)
  AUTHORS   Vickers CE, Bydder SF, Zhou Y, Nielsen LK
  TITLE     Dual gene expression cassette vectors with antibiotic selection 
            markers for engineering in Saccharomyces cerevisiae.
  JOURNAL   Microb Cell Fact. 2013 Oct 25;12(1):96.
  PUBMED    24161108
REFERENCE   2  (bases 1 to 6645)
  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: "2013-10-25"; volume: "12(1)"; pages: "96"
COMMENT     Sequence Label: pCEV-G3-Km
FEATURES             Location/Qualifiers
     source          1..6645
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(2..167)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     88..112
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(315..338)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        560..1001
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    560..677
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     972..995
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     promoter        1018..1424
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     primer_bind     1444..1463
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1444..1462
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1480..1509
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      1539..1728
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1800..1817)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1971..2559)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2051..2070)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2730..3521,3522..3590))
                     /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     3353..3372
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3591..3695)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3722..5064
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(4685..4732)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     protein_bind    5084..5117
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            5172..6528
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        5172..5515
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             5516..6325
                     /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(5583..5602)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      6331..6528
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    6591..6624
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
ORIGIN
        1 ggagcgacct catgctatac ctgagaaagc aacctgacct acaggaaaga gttactcaag
       61 aataagaatt ttcgttttaa aacctaagag tcactttaaa atttgtatac acttattttt
      121 tttataactt atttaataat aaaaatcata aatcataaga aattcgctta tttagaagtg
      181 tcaacaacgt atctaccaac gatttgaccc ttttccatct tttcgtaaat ttctggcaag
      241 gtagacaagc cgacaacctt gattggagac ttgaccaaac ctctggcgaa gaattgttaa
      301 ttaagagctc agatcttatc gtcgtcatcc ttgtaatcca tcgatactag tgaattttca
      361 aaaattctta cttttttttt ggatggacgc aaagaagttt aataatcata ttacatggca
      421 ttaccaccat atacatatcc atatctaatc ttacttatat gttgtggaaa tgtaaagagc
      481 cccattatct tagcctaaaa aaaccttctc tttggaactt tcagtaatac gcttaactgc
      541 tcattgctat attgaagtac ggattagaag ccgccgagcg ggcgacagcc ctccgacgga
      601 agactctcct ccgtgcgtcc tcgtcttcac cggtcgcgtt cctgaaacgc agatgtgcct
      661 cgcgccgcac tgctccgaac aataaagatt ctacaatact agcttttatg gttatgaaga
      721 ggaaaaattg gcagtaacct ggccccacaa accttcaaat taacgaatca aattaacaac
      781 cataggatga taatgcgatt agttttttag ccttatttct ggggtaatta atcagcgaag
      841 cgatgatttt tgatctatta acagatatat aaatggaaaa gctgcataac cactttaact
      901 aatactttca acattttcag tttgtattac ttcttattca aatgtcataa aagtatcaac
      961 aaaaaattgt taatatacct ctatacttta acgtcaagga gaaaaaacgg ccggccgcac
     1021 acaccatagc ttcaaaatgt ttctactcct tttttactct tccagatttt ctcggactcc
     1081 gcgcatcgcc gtaccacttc aaaacaccca agcacagcat actaaatttc ccctctttct
     1141 tcctctaggg tgtcgttaat tacccgtact aaaggtttgg aaaagaaaaa agagaccgcc
     1201 tcgtttcttt ttcttcgtcg aaaaaggcaa taaaaatttt tatcacgttt ctttttcttg
     1261 aaaatttttt tttttgattt ttttctcttt cgatgacctc ccattgatat ttaagttaat
     1321 aaacggtctt caatttctca agtttcagtt tcatttttct tgttctatta caactttttt
     1381 tacttcttgc tcattagaaa gaaagcatag caatctaatc taagttttaa ttacaaggat
     1441 ccgtaatacg actcactata gggcccgggc gtcgacatgg aacagaagtt gatttccgaa
     1501 gaagacctcg agtaagcttg gtaccgcggc tagctaagat ccgctctaac cgaaaaggaa
     1561 ggagttagac aacctgaagt ctaggtccct atttattttt ttatagttat gttagtatta
     1621 agaacgttat ttatatttca aatttttctt ttttttctgt acagacgcgt gtacgcatgt
     1681 aacattatac tgaaaacctt gcttgagaag gttttgggac gctcgaagat ccagctgcat
     1741 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     1801 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     1861 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     1921 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     1981 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     2041 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     2101 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     2161 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2221 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     2281 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     2341 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     2401 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     2461 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     2521 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     2581 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     2641 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     2701 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     2761 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     2821 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     2881 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     2941 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3001 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     3061 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     3121 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     3181 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     3241 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     3301 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     3361 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     3421 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     3481 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     3541 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     3601 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     3661 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     3721 gaacgaagca tctgtgcttc attttgtaga acaaaaatgc aacgcgagag cgctaatttt
     3781 tcaaacaaag aatctgagct gcatttttac agaacagaaa tgcaacgcga aagcgctatt
     3841 ttaccaacga agaatctgtg cttcattttt gtaaaacaaa aatgcaacgc gagagcgcta
     3901 atttttcaaa caaagaatct gagctgcatt tttacagaac agaaatgcaa cgcgagagcg
     3961 ctattttacc aacaaagaat ctatacttct tttttgttct acaaaaatgc atcccgagag
     4021 cgctattttt ctaacaaagc atcttagatt actttttttc tcctttgtgc gctctataat
     4081 gcagtctctt gataactttt tgcactgtag gtccgttaag gttagaagaa ggctactttg
     4141 gtgtctattt tctcttccat aaaaaaagcc tgactccact tcccgcgttt actgattact
     4201 agcgaagctg cgggtgcatt ttttcaagat aaaggcatcc ccgattatat tctataccga
     4261 tgtggattgc gcatactttg tgaacagaaa gtgatagcgt tgatgattct tcattggtca
     4321 gaaaattatg aacggtttct tctattttgt ctctatatac tacgtatagg aaatgtttac
     4381 attttcgtat tgttttcgat tcactctatg aatagttctt actacaattt ttttgtctaa
     4441 agagtaatac tagagataaa cataaaaaat gtagaggtcg agtttagatg caagttcaag
     4501 gagcgaaagg tggatgggta ggttatatag ggatatagca cagagatata tagcaaagag
     4561 atacttttga gcaatgtttg tggaagcggt attcgcaata ttttagtagc tcgttacagt
     4621 ccggtgcgtt tttggttttt tgaaagtgcg tcttcagagc gcttttggtt ttcaaaagcg
     4681 ctctgaagtt cctatacttt ctagagaata ggaacttcgg aataggaact tcaaagcgtt
     4741 tccgaaaacg agcgcttccg aaaatgcaac gcgagctgcg cacatacagc tcactgttca
     4801 cgtcgcacct atatctgcgt gttgcctgta tatatatata catgagaaga acggcatagt
     4861 gcgtgtttat gcttaaatgc gtacttatat gcgtctattt atgtaggatg aaaggtagtc
     4921 tagtacctcc tgtgatatta tcccattcca tgcggggtat cgtatgcttc cttcagcact
     4981 accctttagc tgttctatat gctgccactc ctcaattgga ttagtctcat ccttcaatgc
     5041 tatcatttcc tttgatattg gatcatggta gacaaccctt aatataactt cgtataatgt
     5101 atgctatacg aagttattag gtctagagat ctgtttagct tgcctcgtcc ccgccgggtc
     5161 acccggccag cgacatggag gcccagaata ccctccttga cagtcttgac gtgcgcagct
     5221 caggggcatg atgtgactgt cgcccgtaca tttagcccat acatccccat gtataatcat
     5281 ttgcatccat acattttgat ggccgcacgg cgcgaagcaa aaattacggc tcctcgctgc
     5341 agacctgcga gcagggaaac gctcccctca cagacgcgtt gaattgtccc cacgccgcgc
     5401 ccctgtagag aaatataaaa ggttaggatt tgccactgag gttcttcttt catatacttc
     5461 cttttaaaat cttgctagga tacagttctc acatcacatc cgaacataaa caaccatggg
     5521 taaggaaaag actcacgttt cgaggccgcg attaaattcc aacatggatg ctgatttata
     5581 tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct atcgattgta
     5641 tgggaagccc gatgcgccag agttgtttct gaaacatggc aaaggtagcg ttgccaatga
     5701 tgttacagat gagatggtca gactaaactg gctgacggaa tttatgcctc ttccgaccat
     5761 caagcatttt atccgtactc ctgatgatgc atggttactc accactgcga tccccggcaa
     5821 aacagcattc caggtattag aagaatatcc tgattcaggt gaaaatattg ttgatgcgct
     5881 ggcagtgttc ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt ttaacagcga
     5941 tcgcgtattt cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag
     6001 tgattttgat gacgagcgta atggctggcc tgttgaacaa gtctggaaag aaatgcataa
     6061 gcttttgcca ttctcaccgg attcagtcgt cactcatggt gatttctcac ttgataacct
     6121 tatttttgac gaggggaaat taataggttg tattgatgtt ggacgagtcg gaatcgcaga
     6181 ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc cttcattaca
     6241 gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat tgcagtttca
     6301 tttgatgctc gatgagtttt tctaatcagt actgacaata aaaagattct tgttttcaag
     6361 aacttgtcat ttgtatagtt tttttatatt gtagttgttc tattttaatc aaatgttagc
     6421 gtgatttata ttttttttcg cctcgacatc atctgcccag atgcgaagtt aagtgcgcag
     6481 aaagtaatat catgcgtcaa tcgtatgtga atgctggtcg ctatactgct gtcgattcga
     6541 tactaacgcc gccatccagt gtcgaaaacg agctctcgag aacccttaat ataacttcgt
     6601 ataatgtatg ctatacgaag ttattaggtg atatcagatc cacta
//
