LOCUS       sequence-66304-e        6574 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TEF1 and HXT7 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 6574)
  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 6574)
  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-G4-Km
FEATURES             Location/Qualifiers
     source          1..6574
                     /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        947..1353
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     primer_bind     1373..1392
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1373..1391
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1409..1438
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      1468..1657
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1729..1746)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1900..2488)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1980..1999)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2659..3450,3451..3519))
                     /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     3282..3301
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3520..3624)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3651..4993
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(4614..4661)
                     /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    5013..5046
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            5101..6457
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        5101..5444
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             5445..6254
                     /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(5512..5531)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      6260..6457
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    6520..6553
                     /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 ttttttgatt
      361 aaaattaaaa aaactttttg tttttgtgtt tattctttgt tcttagaaaa gacaagttga
      421 gcttgtttgt tcttgatgtt ttattatttt acaatagctg caaatgaaga atagattcga
      481 acattgtgaa gtattggcat atatcgtctc tatttatact tttttttttt cagttctagt
      541 atattttgta ttttcctcct tttcattctt tcagttgcca ataagttaca ggggatctcg
      601 aaagatggtg gggatttttc cttgaaagac gactttttgc catctaattt ttccttgttg
      661 cctctgaaaa ttatccagca gaagcaaatg taaaagatga acctcagaag aacacgcagg
      721 ggcccgaaat tgttcctacg agaagtagtg ggtcataaaa agtttattcc ctggaaaaaa
      781 aattttgcgt tgcctttctg gagaattttt tcgaattagc gtgctgccac tgcatgcatt
      841 tctgagaagt gtgggcattc ttccaccagt tgttcctcct aaaaaaaaaa agatttccta
      901 ccccgcacaa attcctgcat acccctcatt tccacggggc cggccgcaca caccatagct
      961 tcaaaatgtt tctactcctt ttttactctt ccagattttc tcggactccg cgcatcgccg
     1021 taccacttca aaacacccaa gcacagcata ctaaatttcc cctctttctt cctctagggt
     1081 gtcgttaatt acccgtacta aaggtttgga aaagaaaaaa gagaccgcct cgtttctttt
     1141 tcttcgtcga aaaaggcaat aaaaattttt atcacgtttc tttttcttga aaattttttt
     1201 ttttgatttt tttctctttc gatgacctcc cattgatatt taagttaata aacggtcttc
     1261 aatttctcaa gtttcagttt catttttctt gttctattac aacttttttt acttcttgct
     1321 cattagaaag aaagcatagc aatctaatct aagttttaat tacaaggatc cgtaatacga
     1381 ctcactatag ggcccgggcg tcgacatgga acagaagttg atttccgaag aagacctcga
     1441 gtaagcttgg taccgcggct agctaagatc cgctctaacc gaaaaggaag gagttagaca
     1501 acctgaagtc taggtcccta tttatttttt tatagttatg ttagtattaa gaacgttatt
     1561 tatatttcaa atttttcttt tttttctgta cagacgcgtg tacgcatgta acattatact
     1621 gaaaaccttg cttgagaagg ttttgggacg ctcgaagatc cagctgcatt aatgaatcgg
     1681 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     1741 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     1801 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     1861 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     1921 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     1981 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     2041 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     2101 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     2161 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     2221 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     2281 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     2341 gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     2401 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     2461 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     2521 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     2581 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     2641 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     2701 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     2761 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     2821 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     2881 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     2941 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     3001 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     3061 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     3121 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     3181 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     3241 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     3301 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     3361 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     3421 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     3481 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     3541 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     3601 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg aacgaagcat
     3661 ctgtgcttca ttttgtagaa caaaaatgca acgcgagagc gctaattttt caaacaaaga
     3721 atctgagctg catttttaca gaacagaaat gcaacgcgaa agcgctattt taccaacgaa
     3781 gaatctgtgc ttcatttttg taaaacaaaa atgcaacgcg agagcgctaa tttttcaaac
     3841 aaagaatctg agctgcattt ttacagaaca gaaatgcaac gcgagagcgc tattttacca
     3901 acaaagaatc tatacttctt ttttgttcta caaaaatgca tcccgagagc gctatttttc
     3961 taacaaagca tcttagatta ctttttttct cctttgtgcg ctctataatg cagtctcttg
     4021 ataacttttt gcactgtagg tccgttaagg ttagaagaag gctactttgg tgtctatttt
     4081 ctcttccata aaaaaagcct gactccactt cccgcgttta ctgattacta gcgaagctgc
     4141 gggtgcattt tttcaagata aaggcatccc cgattatatt ctataccgat gtggattgcg
     4201 catactttgt gaacagaaag tgatagcgtt gatgattctt cattggtcag aaaattatga
     4261 acggtttctt ctattttgtc tctatatact acgtatagga aatgtttaca ttttcgtatt
     4321 gttttcgatt cactctatga atagttctta ctacaatttt tttgtctaaa gagtaatact
     4381 agagataaac ataaaaaatg tagaggtcga gtttagatgc aagttcaagg agcgaaaggt
     4441 ggatgggtag gttatatagg gatatagcac agagatatat agcaaagaga tacttttgag
     4501 caatgtttgt ggaagcggta ttcgcaatat tttagtagct cgttacagtc cggtgcgttt
     4561 ttggtttttt gaaagtgcgt cttcagagcg cttttggttt tcaaaagcgc tctgaagttc
     4621 ctatactttc tagagaatag gaacttcgga ataggaactt caaagcgttt ccgaaaacga
     4681 gcgcttccga aaatgcaacg cgagctgcgc acatacagct cactgttcac gtcgcaccta
     4741 tatctgcgtg ttgcctgtat atatatatac atgagaagaa cggcatagtg cgtgtttatg
     4801 cttaaatgcg tacttatatg cgtctattta tgtaggatga aaggtagtct agtacctcct
     4861 gtgatattat cccattccat gcggggtatc gtatgcttcc ttcagcacta ccctttagct
     4921 gttctatatg ctgccactcc tcaattggat tagtctcatc cttcaatgct atcatttcct
     4981 ttgatattgg atcatggtag acaaccctta atataacttc gtataatgta tgctatacga
     5041 agttattagg tctagagatc tgtttagctt gcctcgtccc cgccgggtca cccggccagc
     5101 gacatggagg cccagaatac cctccttgac agtcttgacg tgcgcagctc aggggcatga
     5161 tgtgactgtc gcccgtacat ttagcccata catccccatg tataatcatt tgcatccata
     5221 cattttgatg gccgcacggc gcgaagcaaa aattacggct cctcgctgca gacctgcgag
     5281 cagggaaacg ctcccctcac agacgcgttg aattgtcccc acgccgcgcc cctgtagaga
     5341 aatataaaag gttaggattt gccactgagg ttcttctttc atatacttcc ttttaaaatc
     5401 ttgctaggat acagttctca catcacatcc gaacataaac aaccatgggt aaggaaaaga
     5461 ctcacgtttc gaggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat
     5521 gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgattgtat gggaagcccg
     5581 atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg
     5641 agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta
     5701 tccgtactcc tgatgatgca tggttactca ccactgcgat ccccggcaaa acagcattcc
     5761 aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc
     5821 tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat cgcgtatttc
     5881 gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg
     5941 acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataag cttttgccat
     6001 tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg
     6061 aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg
     6121 atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt
     6181 ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg
     6241 atgagttttt ctaatcagta ctgacaataa aaagattctt gttttcaaga acttgtcatt
     6301 tgtatagttt ttttatattg tagttgttct attttaatca aatgttagcg tgatttatat
     6361 tttttttcgc ctcgacatca tctgcccaga tgcgaagtta agtgcgcaga aagtaatatc
     6421 atgcgtcaat cgtatgtgaa tgctggtcgc tatactgctg tcgattcgat actaacgccg
     6481 ccatccagtg tcgaaaacga gctctcgaga acccttaata taacttcgta taatgtatgc
     6541 tatacgaagt tattaggtga tatcagatcc acta
//
