LOCUS       sequence-116407-        6959 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EasyClone system-based yeast integrative vector carrying 
            loxP-flanked KlLEU2 marker, integration into S. cerevisiae XII-2 
            chromosomal location, USER site for cloning, amp resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6959)
  AUTHORS   Stovicek V, Borja GM, Forster J, Borodina I
  TITLE     EasyClone 2.0: expanded toolkit of integrative vectors for stable 
            gene expression in industrial Saccharomyces cerevisiae strains.
  JOURNAL   J Ind Microbiol Biotechnol. 2015 Sep 16.
  PUBMED    26376869
REFERENCE   2  (bases 1 to 6959)
  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: "J Ind 
            Microbiol Biotechnol. 2015 Sep 16."
COMMENT     Sequence Label: pCfB2192
FEATURES             Location/Qualifiers
     source          1..6959
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      30..219
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     protein_bind    267..300
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2589..2622
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(2665..2684)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2666..2684)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2694..2710)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2703..2725)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     protein_bind    complement(3372..3405)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(3875..3891)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3990..4007)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4161..4749)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4241..4260)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4873..5664,5665..5733))
                     /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     5496..5515
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5734..5825)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5867..5886
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5892..5911
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     protein_bind    6118..6151
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     terminator      complement(6765..6930)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     6851..6875
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
ORIGIN
        1 caacggaatg cgtgcgatcg cgtgcattca tccgctctaa ccgaaaagga aggagttaga
       61 caacctgaag tctaggtccc tatttatttt tttatagtta tgttagtatt aagaacgtta
      121 tttatatttc aaatttttct tttttttctg tacagacgcg tgtacgcatg taacattata
      181 ctgaaaacct tgcttgagaa ggttttggga cgctcgaaga tcgcgtcagc tgaagcttcg
      241 tacgctgcag gtcgacaacc cttaatataa cttcgtataa tgtatgctat acgaagttat
      301 taggtctaga gatccgcagg ctaaccggaa cctgtattat ttagtttatg ctacgttaaa
      361 taaagacctt tcgttcacat aactgaatgt gtaatggcct tgagatttca agcataccaa
      421 gttggtggag acggggtcgt tacaaaagac tctttaagcc aagatttcct tgacagcctt
      481 ggcgatagca tcgccaacct cagtggtaga gttagaacca ccaaggtcac cggttctgac
      541 acctgcatcc aagacatttc taacagcttc ttcaagagcc ctaccttctt caaccaaatc
      601 caaggataac ttcaacatca tagctgcaga taagatggta gcaattgggt taaccttgtt
      661 tgctggtaaa tctggggcag aaccatgaca tggttcgtac aaaccgaatg ccttgttagt
      721 gtcaggtagg gaagctagag atgcagaagg taataaaccc aaagaacctg gaataacaga
      781 ggcttcatcg gagataatat caccaaacat gttgttggta ataacaacac cgtttagctt
      841 agttggtgat ttaaccaaaa tcatagcagc agagtcgatc aattggtgct gaacagttaa
      901 ttgtgggaac tcagtcttga tggtttcttc aacagtcttt ctccacaatc tggaagaggc
      961 aagcacgtta gccttgtcaa gtgaccagat tggtaatggt gggttttgtt gcaatgccaa
     1021 gaaagcagcc attcttgtaa ttctttgaac ttcaggaaca ctgtatttct cagagtccca
     1081 agcaactccg tcaccttcat cttcttttct ttcaccaaag tagataccac caaccaattc
     1141 tctaacgacg acgaaatcgg taccctttgc atattcaggc ttcaaaggag aaagatctag
     1201 taaagaatca gaagcaaagt tacatggtct taagttggcg tatagaccca attcctttct
     1261 gatcttcaat agaccttgtt ctggtctaac tgcgcccgta ccccattttg gaccaccaac
     1321 agcacctagt aagacagcat cggctttctt agaggcttct agagcttcat ctggtaaagg
     1381 agtgccagtg gcatcgatgg cagcaccccc gatcaagtga tgttggaaat tgaacttaat
     1441 ttctggacgg acttcagcaa tggcattcaa gaccttaata gcttcgtcag taacttcttt
     1501 accgacgtga tcacccggta ggacaacgat attcttagac atggtataat ctgtgtagtg
     1561 tgggatactt tttacttctt caaataggta tcaacaataa aaataaatcg aagcaaatgt
     1621 aggaatgcgt taaagcagat gtacttactt agagtacata gatatattta tataattcaa
     1681 tatataaaag tatatgaagc atctgatgtt gaacctgtca tgactctaaa ctggaatggg
     1741 tagttatgct tctgaaggtt ttccgttaga aaatgagtgc agggatcagg cccggaaccg
     1801 gttttagcct gaaaggaaaa gaaaaaaatg cggtgattac tagcacgtga ctgcgctgaa
     1861 ttggaatcca ccacaagtac agcgaatggc aagagggaag gggaatatat tgtgcctggc
     1921 ggtagtctgc tatatcaaat gttaaacata actaaatgca gtagtgaata gagtaatgta
     1981 tttctacata tgtgggtagg gccaatggga gcccgatgtc tattgtacag atattttcct
     2041 tttatagtta atcaattcaa cttggaaatc attgagtcga tatctacttg gataactgga
     2101 gccgtggcag cgtttgaaat ttgacgagtc tccttgtttt gttcgcgcat tttcagttta
     2161 ttcaaacggg agatggcacc gaatccactg acttctctgt tcaatttgaa tggtaatagg
     2221 aagaaagtac atttcgagaa attttcgccc aacgtcgaaa tctcatggat gatatcctcg
     2281 atacaaatga caccttcatc acctaatttc tcttcaacga tattgttgtc attcaagatt
     2341 atctcttttg gttctttatc ttctggcctt tgccacatta ttctagatct cttttggatt
     2401 aaagaacgga tagatgcaag tgaaggttgt ccgataacaa tatatggtga tacaagtttc
     2461 aacaatgggt acacgtcctt ggtaagtttg atgaaaacac cagtgttcaa ctcaacaagt
     2521 ctcaaaagag ataacacttt gtaagccttt gctgggatct tcacagcgag ctctcgagaa
     2581 cccttaatat aacttcgtat aatgtatgct atacgaagtt attaggtgat atcagatcca
     2641 ctagtggcct atgcacccaa ttcgccctat agtgagtcgt attacgcgcg ctcactggcc
     2701 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccctgcagg actagtgctg
     2761 aggcattaat ctactatcgg cgactctctc gaaatttttc ttaacgcgtc cttgtactgc
     2821 gtctaacgct tttgccactt ggatttctat tataggaaat agtctcactt actgggcgac
     2881 gaattttcgc gttttgatga agcacaggaa gaatttcttt tttttttggc ttcttctggt
     2941 tccgtttttt acgcgcacaa atctaaaaaa agaaataatt ataacctagt ctcgaaaatt
     3001 ttcatcgatc cattcgttcc tttttttcga ttttttcaga tcaaaattct tgtttctttc
     3061 tttgtcttag tttatattaa aagatatttt gattttactc ctgaactatt tattctttct
     3121 aagaaggcca gaacactaca gctgttttaa ccgactacga agttctccat tctcgaacac
     3181 tagccttcat ttaccaaaca ggaactagcg tatatcatta gtccttattc gaaaagagat
     3241 tggtagatat ttattgtagt ttgtgagaag gagaaaatac tgtcattgga ctgatagtta
     3301 gaggacatta acctctctta cgttcgctca ttaagacctc agcgcggccg caaatttaaa
     3361 taaaatgaag tgaagttcct atactttcta gagaatagga acttctatag tgagtcgaat
     3421 aagggcgaca caaaatttat tctaaatgca taataaatac tgataacatc ttatagtttg
     3481 tattatattt tgtattatcg ttgacatgta taattttgat atcaaaaact gattttccct
     3541 ttattatttt cgagatttat tttcttaatt ctctttaaca aactagaaat attgtatata
     3601 caaaaaatca taaataatag atgaatagtt taattatagg tgttcatcaa tcgaaaaagc
     3661 aacgtatctt atttaaagtg cgttgctttt ttctcattta taaggttaaa taattctcat
     3721 atatcaagca aagtgacagg cgcccttaaa tattctgaca aatgctcttt ccctaaactc
     3781 cccccataaa aaaacccgcc gaagcgggtt tttacgttat ttgcggatta acgattactc
     3841 gttatcagaa ccgcccaggg ggcccgagct taagactggc cgtcgtttta caacacagaa
     3901 agagtttgta gaaacgcaaa aaggccatcc gtcaggggcc ttctgcttag tttgatgcct
     3961 ggcagttccc tactctcgcc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     4021 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     4081 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     4141 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     4201 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     4261 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     4321 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     4381 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     4441 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     4501 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     4561 gagttcttga agtggtgggc taactacggc tacactagaa gaacagtatt tggtatctgc
     4621 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     4681 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     4741 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgacgcg
     4801 cgcgtaactc acgttaaggg attttggtca tgagcttgcg ccgtcccgtc aagtcagcgt
     4861 aatgctctgc ttttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     4921 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     4981 accatctggc cccagcgctg cgatgatacc gcgagaacca cgctcaccgg ctccggattt
     5041 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5101 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5161 tagtttgcgc aacgttgttg ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5221 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5281 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     5341 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     5401 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     5461 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     5521 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     5581 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     5641 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     5701 aataagggcg acacggaaat gttgaatact catattcttc ctttttcaat attattgaag
     5761 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     5821 acaaataggg gtcagtgtta caaccaatta accaattctg aacattatcg cgagcccatt
     5881 tatacctgaa tatggctcat aacacccctt gtttgcctgg cggcagtagc gcggtggtcc
     5941 cacctgaccc catgccgaac tcagaagtga aacgccgtag cgccgatggt agtgtgggga
     6001 ctccccatgc gagagtaggg aactgccagg catcaaataa aacgaaaggc tcagtcgaaa
     6061 gactgggcct ttcgcccggg ctaattatgg ggtgtcgccc ttattcgact ctatagtgaa
     6121 gttcctattc tctagaaagt ataggaactt ctgaagtggg gatttaaatg cggccgcgct
     6181 gagggtttaa tcggcatgca aacatctaca caattagcaa gggcaatcca tattttgtct
     6241 tttcgcgccc tggaaaggcc taagtaatgt cgtaaacgca ttctatctgt acttcaactc
     6301 tcctctgtgc attggtttgt gcaaatcaca ttttacgata ctgccagata tatgcaaaaa
     6361 gagaaaacca agggaccaga acaaagcaaa attacgatat tcttcgaatt ccttcgtgct
     6421 tgactaagac aaagggatgg acgtagcgat ttttagcggg ccaagaactg gttccgaaaa
     6481 agcacaggta caccgaaccc tcagctaagg agggacagca ccgatgcgga aggacaaact
     6541 ttctttttgc ctatcacagt atcttatcga gctaactatt ttcgacacac atgaaaaagc
     6601 agaaatatta acgaaaaaga aaagaaagac catgtcatgt acgggcaatc agaatctgta
     6661 acaagcgcca tttttttttc tgtatcgggc cctccttact gctctccttc cgtgtaacgc
     6721 gttatgatta agtcctcagc gagctcgcat ggaatgcgtg cgatgagcga cctcatgcta
     6781 tacctgagaa agcaacctga cctacaggaa agagttactc aagaataaga attttcgttt
     6841 taaaacctaa gagtcacttt aaaatttgta tacacttatt ttttttataa cttatttaat
     6901 aataaaaatc ataaatcata agaaattcgc ttatttagaa gtgtcaacaa cgtatctac
//
