LOCUS       sequence-148873-        6269 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCfB2991 is a vector for multiple integrations at sites sharing 
            homology with Ty1Cons2. The selective marker is Kl.LEU2. 
            Additionally it contains a USER cassette..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6269)
  AUTHORS   Maury J, Germann SM, Baallal Jacobsen SA, Jensen NB, Kildegaard KR, 
            Herrgard MJ, Schneider K, Koza A, Forster J, Nielsen J, Borodina I
  TITLE     EasyCloneMulti: A Set of Vectors for Simultaneous and Multiple 
            Genomic Integrations in Saccharomyces cerevisiae.
  JOURNAL   PLoS One. 2016 Mar 2;11(3):e0150394. doi: 
            10.1371/journal.pone.0150394. eCollection 2016.
  PUBMED    26934490
REFERENCE   2  (bases 1 to 6269)
  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: "PLoS One.";
            date: "2016-03-2"; volume: "11(3):e0150394. doi"; pages: " 
            10.1371/journal.pone.0150394. eCollection 2016"
COMMENT     Sequence Label: pCfB2991
FEATURES             Location/Qualifiers
     source          1..6269
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      20..209
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     protein_bind    257..290
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2607..2640
                     /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(2683..2702)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2684..2702)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2712..2728)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2721..2743)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     protein_bind    complement(3002..3035)
                     /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(3505..3521)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3620..3637)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3791..4379)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3871..3890)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4503..5294,5295..5363))
                     /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     5126..5145
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5364..5455)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5497..5516
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5522..5541
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     protein_bind    5748..5781
                     /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(6065..6230)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     6151..6175
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
ORIGIN
        1 cgtgcgatcg cgtgcattca tccgctctaa ccgaaaagga aggagttaga caacctgaag
       61 tctaggtccc tatttatttt tttatagtta tgttagtatt aagaacgtta tttatatttc
      121 aaatttttct tttttttctg tacagacgcg tgtacgcatg taacattata ctgaaaacct
      181 tgcttgagaa ggttttggga cgctcgaaga tcgcgtcagc tgaagcttcg tacgctgcag
      241 gtcgacaacc cttaatataa cttcgtataa tgtatgctat acgaagttat taggtctaga
      301 gatcccaata caacagatca cgtgatcttt ttgtaagatg aagttgaagt agtgttgcac
      361 cgtgccaatg caggtggcta ttagattaaa tatgtgattt gttctattaa gtttcctgta
      421 taattacaaa tgaataacga aatgagacaa agaagagaac caatttttac aagcagccaa
      481 gatttccttg acagccttgg cgatagcatc gccaacctca gtggtagagt tagaaccacc
      541 aaggtcaccg gttctgacac ctgcatccaa gacatttcta acagcttctt caagagccct
      601 accttcttca accaaatcca aggataactt caacatcata gctgcagata agatggtagc
      661 aattgggtta accttgtttg ctggtaaatc tggggcagaa ccatgacatg gttcgtacaa
      721 accgaatgcc ttgttagtgt caggtaggga agctagagat gcagaaggta ataaacccaa
      781 agaacctgga ataacagagg cttcatcgga gataatatca ccaaacatgt tgttggtaat
      841 aacaacaccg tttagcttag ttggtgattt aaccaaaatc atagcagcag agtcgatcaa
      901 ttggtgctga acagttaatt gtgggaactc agtcttgatg gtttcttcaa cagtctttct
      961 ccacaatctg gaagaggcaa gcacgttagc cttgtcaagt gaccagattg gtaatggtgg
     1021 gttttgttgc aatgccaaga aagcagccat tcttgtaatt ctttgaactt caggaacact
     1081 gtatttctca gagtcccaag caactccgtc accttcatct tcttttcttt caccaaagta
     1141 gataccacca accaattctc taacgacgac gaaatcggta ccctttgcat attcaggctt
     1201 caaaggagaa agatctagta aagaatcaga agcaaagtta catggtctta agttggcgta
     1261 tagacccaat tcctttctga tcttcaatag accttgttct ggtctaactg cgcccgtacc
     1321 ccattttgga ccaccaacag cacctagtaa gacagcatcg gctttcttag aggcttctag
     1381 agcttcatct ggtaaaggag tgccagtggc atcgatggca gcacccccga tcaagtgatg
     1441 ttggaaattg aacttaattt ctggacggac ttcagcaatg gcattcaaga ccttaatagc
     1501 ttcgtcagta acttctttac cgacgtgatc acccggtagg acaacgatat tcttagacat
     1561 ggtataatct gtgtagtgtg ggatactttt tacttcttca aataggtatc aacaataaaa
     1621 ataaatcgaa gcaaatgtag gaatgcgtta aagcagatgt acttacttag agtacataga
     1681 tatatttata taattcaata tataaaagta tatgaagcat ctgatgttga acctgtcatg
     1741 actctaaact ggaatgggta gttatgcttc tgaaggtttt ccgttagaaa atgagtgcag
     1801 ggatcaggcc cggaaccggt tttagcctga aaggaaaaga aaaaaatgcg gtgattacta
     1861 gcacgtgact gcgctgaatt ggaatccacc acaagtacag cgaatggcaa gagggaaggg
     1921 gaatatattg tgcctggcgg tagtctgcta tatcaaatgt taaacataac taaatgcagt
     1981 agtgaataga gtaatgtatt tctacatatg tgggtagggc caatgggagc ccgatgtcta
     2041 ttgtacagat attttccttt tatagttaat caattcaact tggaaatcat tgagtcgata
     2101 tctacttgga taactggagc cgtggcagcg tttgaaattt gacgagtctc cttgttttgt
     2161 tcgcgcattt tcagtttatt caaacgggag atggcaccga atccactgac ttctctgttc
     2221 aatttgaatg gtaataggaa gaaagtacat ttcgagaaat tttcgcccaa cgtcgaaatc
     2281 tcatggatga tatcctcgat acaaatgaca ccttcatcac ctaatttctc ttcaacgata
     2341 ttgttgtcat tcaagattat ctcttttggt tctttatctt ctggcctttg ccacattatt
     2401 ctagatctct tttggattaa agaacggata gatgcaagtg aaggttgtcc gataacaata
     2461 tatggtgata caagtttcaa caatgggtac acgtccttgg taagtttgat gaaaacacca
     2521 gtgttcaact caacaagtct caaaagagat aacactttgt aagcctttgc tgggatcttc
     2581 acagcgagct ctcgagaacc cttaatataa cttcgtataa tgtatgctat acgaagttat
     2641 taggtgatat cagatccact agtggcctat gcacccaatt cgccctatag tgagtcgtat
     2701 tacgcgcgct cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     2761 cctgcaggac agacgtcatt agtgctgagg cattaattga tcataaaacg gaatgatgaa
     2821 taatatttat agaattgtgt agaattgcag attccctttt atggattcct aaatcctcga
     2881 ggagaacttc tagtatattc tgtataccta atattatagc ctttatcaac aatggaatcc
     2941 caacaattat ctcaaaattc acatatttct caagatctgc ggccgcactc agacctgaag
     3001 tgaagttcct atactttcta gagaatagga acttctatag tgagtcgaat aagggcgaca
     3061 caaaatttat tctaaatgca taataaatac tgataacatc ttatagtttg tattatattt
     3121 tgtattatcg ttgacatgta taattttgat atcaaaaact gattttccct ttattatttt
     3181 cgagatttat tttcttaatt ctctttaaca aactagaaat attgtatata caaaaaatca
     3241 taaataatag atgaatagtt taattatagg tgttcatcaa tcgaaaaagc aacgtatctt
     3301 atttaaagtg cgttgctttt ttctcattta taaggttaaa taattctcat atatcaagca
     3361 aagtgacagg cgcccttaaa tattctgaca aatgctcttt ccctaaactc cccccataaa
     3421 aaaacccgcc gaagcgggtt tttacgttat ttgcggatta acgattactc gttatcagaa
     3481 ccgcccaggg ggcccgagct taagactggc cgtcgtttta caacacagaa agagtttgta
     3541 gaaacgcaaa aaggccatcc gtcaggggcc ttctgcttag tttgatgcct ggcagttccc
     3601 tactctcgcc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3661 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3721 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3781 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3841 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3901 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3961 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     4021 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     4081 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4141 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4201 agtggtgggc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     4261 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4321 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4381 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgacgcg cgcgtaactc
     4441 acgttaaggg attttggtca tgagcttgcg ccgtcccgtc aagtcagcgt aatgctctgc
     4501 ttttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     4561 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     4621 cccagcgctg cgatgatacc gcgagaacca cgctcaccgg ctccggattt atcagcaata
     4681 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     4741 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     4801 aacgttgttg ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     4861 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     4921 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     4981 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     5041 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     5101 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     5161 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     5221 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     5281 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     5341 acacggaaat gttgaatact catattcttc ctttttcaat attattgaag catttatcag
     5401 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     5461 gtcagtgtta caaccaatta accaattctg aacattatcg cgagcccatt tatacctgaa
     5521 tatggctcat aacacccctt gtttgcctgg cggcagtagc gcggtggtcc cacctgaccc
     5581 catgccgaac tcagaagtga aacgccgtag cgccgatggt agtgtgggga ctccccatgc
     5641 gagagtaggg aactgccagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct
     5701 ttcgcccggg ctaattatgg ggtgtcgccc ttattcgact ctatagtgaa gttcctattc
     5761 tctagaaagt ataggaactt ctgaagtggg gatttaaagt cggtgtccgc gctgagggtt
     5821 taatggcgcg ccgcggccgc ccgcggtgtt ggaataaaaa tcaactatca tctactaact
     5881 agtatttacg ttactagtat attatcatat acggtgttag aagatgacgc aaatgatgag
     5941 aaatagtcat ctaaattagt ggaagctgaa acgcaaggat tgataatgta ataggatcaa
     6001 tgaatattaa catagctagc attaagtcct cagcgagcac gctgcttcat ggaatgcgtg
     6061 cgatgagcga cctcatgcta tacctgagaa agcaacctga cctacaggaa agagttactc
     6121 aagaataaga attttcgttt taaaacctaa gagtcacttt aaaatttgta tacacttatt
     6181 ttttttataa cttatttaat aataaaaatc ataaatcata agaaattcgc ttatttagaa
     6241 gtgtcaacaa cgtatctacc aacggaatg
//
